Thermal treatment of acne with coated metal nanoparticles

ABSTRACT

Treatment of skin tissue with photoactive materials and light, such as nanoparticles and formulations which are useful for cosmetic, diagnostic and therapeutic applications to mammals such as humans. In particular, embodiments of thermal treatment of acne with coated metal nanoparticles are disclosed.

CROSS REFERENCE AND INCORPORATION BY REFERENCE TO ANY PRIORITYAPPLICATIONS

This application is a continuation of U.S. patent application Ser. No.14/020,423 filed Sep. 6, 2013, which is a continuation of U.S. patentapplication Ser. No. 13/219,514 filed Aug. 26, 2011, which claims thebenefit of priority under 35 U.S.C. §119(e) of U.S. ProvisionalApplication Nos. 61/402,305 filed Aug. 27, 2010; 61/422,612 filed Dec.13, 2010, and 61/516,308 filed Apr. 1, 2011; each of which is herebyincorporated by reference in its entirety. Any and all applications forwhich a foreign or domestic priority claim is identified in theApplication Data Sheet as filed with the present application are herebyincorporated by reference under 37 CFR 1.57.

PARTIES OF JOINT RESEARCH AGREEMENT

The invention described herein was created subject to a Joint ResearchAgreement between Sienna Labs, Inc. and Nanocomposix, Inc.

BACKGROUND

1. Field of the Invention

The field of the invention is nanoparticles and/or photoactive compoundsfor use in cosmetic, diagnostic and/or therapeutic procedures.

2. Description of the Related Art

Laser treatments of the skin are widely known and have been highlytouted for therapeutic and cosmetic utility. Therapeutically, potentialuses for laser skin therapy include laser ablation of cancerous cells incancer patients and laser ablation of damaged tissue in burn victims.Cosmetic applications for laser skin therapy are much more numerous, andinclude hair removal/reduction, treatment of dyschromia, shrinking ofthe skin following operations such as liposuction, acne treatment,chemical or physical abrasion of unwanted markings on the skin, surgicaltreatments including nose reduction and face- and neck-lifts, and otheraesthetic skin remodeling purposes.

SUMMARY

Despite the promise of laser therapy for skin therapeutics andcosmetics, current laser procedures have limited efficacy, requiringprohibitive numbers of repeated treatments and driving increased costs.Suboptimal laser treatments also have limited specificity, resulting indebilitating clinical side effects, such as non-specific skin damage,skin irritation and scarring.

Light-based hair removal systems suffer from particularly low rates ofefficacy at removing light hair (vellus, blonde, gray, red hair).Multiple (even 6 or more) treatments are insufficient to achieve atherapeutic result in blonde- gray- or red-haired patients, even withthe use of topically applied chromophores such as carbon. In addition tolight hair removal, thermoablative technology has untapped potential inthe fields of wound healing, tissue remodeling, vascular repair, andacne treatment.

Acne vulgaris results from obstruction of the pilosebaceous unit,consisting of the hair shaft, hair follicle, sebaceous gland and erectorpili muscle, which leads to accumulation of sebum oil produced from thesebaceous gland and the subsequent colonization of bacteria within thefollicle. Microcomedones formed as a result of accumulated sebumprogress to non-inflamed skin blemishes (white/blackheads), or to skinblemishes which recruit inflammatory cells and lead to the formation ofpapules, nodules and pus-filled cysts. The sequelae of untreated acnevulgaris often include hyperpigmentation, scarring and disfiguration, aswell as significant psychological distress. Therefore, acne treatmentsseek broadly to reduce the accumulation of sebum and microorganismswithin follicles and the sebaceous gland.

Methods involving light and lasers are promising for the treatment skindisorders, but are still insufficiently effective. Ultraviolet (UV)/bluelight is approved by the FDA for the treatment of mild to moderate acneonly, due to its anti-inflammatory effects mediated on skin cells(keratinocytes), potentially through the action of endogenous porphyrinphotosensitizers within follicles. Exogenous porphirin precursors suchas 5-aminoluveulinic acid (5-ALA) have been formulated for topical ororal delivery and shown to accumulate within sebaceous follicles, absorbphotons from red light exposure and form reactive oxygen species thatdirectly damage cellular membranes and proteins. This procedurecombining porphyrin application and high intensity red light, termed‘photodynamic therapy’, has been demonstrated to reduce sebum productionand acne by 50% for 20 weeks post-irradiation. However, high intensityenergies (50-150 J/cm²) are required to damage sebaceous gland skinstructures, and transdermal porphyrin penetration leads to off-targetside-effects which include sensitivity to light, pain, inflammation,hyper/hypo-pigmentation, and permanent scarring.

For laser therapy to achieve its full utility in the treatment of humanskin disorders, methods to locally induce photo-destruction in skinstructures without affecting surrounding tissues must be achieved.

Provided herein, in certain embodiments, are new compositions andmethods useful in the targeted thermomodulation of target cellpopulations and target tissues, for the purposes of cosmetic treatmentsand the treatment and prevention of chronic and acute diseases anddisorders.

In one aspect, described herein are compositions of matter. For example,in one embodiment, provided is a composition comprising a cosmeticallyacceptable carrier and a plurality of plasmonic nanoparticles in anamount effective to induce thermomodulation in a target tissue regionwith which the composition is topically contacted.

In some embodiments, the composition comprises plasmonic nanoparticlesthat are activated by exposure to energy delivered from a nonlinearexcitation surface plasmon resonance source to the target tissue region.In further or additional embodiments, described herein are compositionscomprising at least one plasmonic nanoparticle that comprises a metal,metallic composite, metal oxide, metallic salt, electric conductor,electric superconductor, electric semiconductor, dielectric, quantum dotor composite from a combination thereof. In further or additionalembodiments, provided herein is a composition wherein a substantialamount of the plasmonic particles present in the composition comprisegeometrically-tuned nanostructures. In certain embodiments, providedherein is a composition wherein plasmonic particles comprise anygeometric shape currently known or to be created that absorb light andgenerate plasmon resonance at a desired wavelength, includingnanoplates, solid nanoshells, hollow nanoshells, nanorods, nanorice,nanospheres, nanofibers, nanowires, nanopyramids, nanoprisms, nanostarsor a combination thereof. In yet additional embodiments, describedherein is a composition wherein the plasmonic particles comprise silver,gold, nickel, copper, titanium, silicon, galadium, palladium, platinum,or chromium.

In some embodiments, provided herein is a composition comprising acosmetically acceptable carrier that comprises an additive, a colorant,an emulsifier, a fragrance, a humectant, a polymerizable monomer, astabilizer, a solvent, or a surfactant. In one embodiment, providedherein is a composition wherein the surfactant is selected from thegroup consisting of: sodium laureth 2-sulfate, sodium dodecyl sulfate,ammonium lauryl sulfate, sodium octech-1/deceth-1 sulfate, lipids,proteins, peptides or derivatives thereof. In one embodiment, providedis a composition wherein a surfactant is present in an amount betweenabout 0.1 and about 10.0% weight-to-weight of the carrier. In yetanother embodiment, the solvent is selected from the group consisting ofwater, propylene glycol, alcohol, hydrocarbon, chloroform, acid, base,acetone, diethyl-ether, dimethyl sulfoxide, dimethylformamide,acetonitrile, tetrahydrofuran, dichloromethane, and ethylacetate. In oneembodiment, the composition comprises plasmonic particles that have anoptical density of at least about 1 O.D. at one or more peak resonancewavelengths.

In further or additional embodiments, described herein is a compositionwherein plasmonic particles comprise a hydrophilic or aliphatic coating,wherein the coating does not substantially adsorb to skin of a mammaliansubject, and wherein the coating comprises polyethylene glycol, silica,silica-oxide, polyvinylpyrrolidone, polystyrene, a protein or a peptide.In yet an additional embodiment, the thermomodulation comprises damage,ablation, thermoablation, lysis, denaturation, deactivation, activation,induction of inflammation, activation of heat shock proteins,perturbation of cell-signaling or disruption to the cellmicroenvironment in the target tissue region. Still further, in certainpresentations the target tissue region comprises a sebaceous gland, acomponent of a sebaceous gland, a sebocyte, a component of a sebocyte,sebum, or hair follicle infundibulum. In further embodiments, the targettissue region comprises a bulge, a bulb, a stem cell, a stem cell niche,a dermal papilla, a cortex, a cuticle, a hair sheath, a medulla, apylori muscle, a Huxley layer, or a Henle layer.

In another aspect, described herein are methods of performing targetedablation of tissue. For example, in one embodiment, provided is a methodfor performing targeted ablation of a tissue to treat a mammaliansubject in need thereof, comprising the steps of i) topicallyadministering to a skin surface of the subject the composition of claim1; ii) providing penetration means to redistribute the plasmonicparticles from the skin surface to a component of dermal tissue; andiii) causing irradiation of the skin surface by light. In further oradditional embodiments, provided is a method wherein the light sourcecomprises excitation of mercury, xenon, deuterium, or a metal-halide,phosphorescence, incandescence, luminescence, light emitting diode, orsunlight. In still further or additional embodiments, provided is amethod wherein the penetration means comprises high frequencyultrasound, low frequency ultrasound, massage, iontophoresis, highpressure air flow, high pressure liquid flow, vacuum, pre-treatment withfractionated photothermolysis or dermabrasion, or a combination thereof.In still further embodiments, provided is a method wherein theirradiation comprises light having a wavelength of light between about200 nm and about 10,000 nm, a fluence of about 1 to about 100joules/cm², a pulse width of about 1 femptosecond to about 1 second, anda repetition frequency of about 1 Hz to about 1 THz.

In a further aspect, provided herein is a composition comprising acosmetically acceptable carrier, an effective amount of sodium dodecylsulfate, and a plurality of plasmonic nanoparticles in an amounteffective to induce thermal damage in a target tissue region with whichthe composition is topically contacted, wherein the nanoparticles havean optical density of at least about 1 O.D. at a resonance wavelength ofabout 810 nanometers or 1064 nanometers, wherein the plasmonic particlescomprise a silica coating from about 5 to about 35 nanometers, whereinthe acceptable carrier comprises water and propylene glycol.

In yet another aspect, provided is a system for laser ablation of hairor treatment of acne comprising a composition and a source of plasmonicenergy suitable for application to the human skin.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is illustrative of schematics depicting certain embodiments ofthe use of formulations for hair removal and acne treatment. Depicted is(A) for hair removal, the plasmonic nanoparticle formulation (black)is 1. applied topically to human skin, 2. delivered deep into thefollicle and washed from the skin surface, 3. irradiated with a clinicallaser at a wavelength resonant to the peak absorption wavelength of theplasmonic particle, and 4. shed from the follicle along with the damagedhair follicle; and (B) for acne treatment, the plasmonic nanoparticleformulation (black) is 1. applied topically to human skin, 2. deliveredspecifically into the sebaceous gland and washed from the skin surface,3. irradiated with a clinical laser at a wavelength resonant to the peakabsorption wavelength of the plasmonic particle, and 4. shed from thetarget site where the accumulated sebum and sebum-producing capabilitiesof the sebaceous gland are destroyed.

FIG. 2 is illustrative of a temperature profile of certain embodimentsof the formulations of plasmonic nanoparticles (SL-001, triangles)provided herein compared to exemplary current clinical dyes carbonlotion (circles), meladine spray (diamonds), and indocyanine green(squares), after exposure to 1064 nm, 20 J/cm², 55 ms laser pulses.SL-001 and dyes were equally diluted at 1:1000 from clinicalconcentration (SL-001 1000 O.D., carbon 20-200 mg/ml, meladine 1 mg/ml,ICG 5 mg/ml). n=3, error S.D. of mean.

FIG. 3 is illustrative of hair follicle penetration offluorescently-labeled nanoparticles determined using porcine skinexplants and confocal imaging of certain embodiments of the subjectmatter described herein. Depicted is (A) schematic of treated porcineskin, sectioned and imaged at an angle to the follicle, in two serial 60μm planes: ‘plane 1’ (showing follicle infundibulum) and ‘plane 2’(showing deep follicle); (B) representative confocal images show redfluorescent nanoparticles (548 nm) within superficial and deep follicle,but not in underlying dermis; and (C) red fluorescent nanoparticlesretained in the deep follicle (˜400 μm) at high magnification. Green istissue autofluorescence.

FIG. 4 is illustrative of a hair follicle penetration of plasmonicnanoparticles determined using porcine skin explants and dark fieldimaging. Shown is (A) schematic of treated porcine skin, sectioned andimaged horizontal to the follicle; (B) bright blue plasmonic particlesare visible in a 1.2 mm deep section, and are differentiated from (C)untreated (negative control) porcine skin, where no pigments arevisible.

FIG. 5 depicts clinical observations in live human skin treated withLaser Only (left forearm) or Plasmonic Particles+Laser (right forearm)demonstrates non-specific and specific photothermal damage. (A,B) In thetop panel, human skin was irradiated with 810 nm laser pulses (30 J/cm2,30 ms, 2 passes) alone (A), or after treatment with a formulation of 830nm resonant, Uncoated plasmonic nanoparticles in 20% propylene glycol(B). The plasmonic nanoparticle formulation was applied with 3 minutemassage, and the skin surface wiped with 3 applications of alternativewater and ethanol before laser irradiation. At 30 minutes followinglaser irradiation, non-specific clinical burns were observed in Bcompared to A, due to significant photothermal heating of residual,Uncoated particles on the skin surface. (C,D) In the bottom panel, humanskin was irradiated with 1064 nm laser pulses (40 J/cm2, 55 ms, 3passes) alone (C), or after treatment with a formulation of 1020 nmresonant, Silica-coated plasmonic nanoparticles in 20% propylene glycol(D). The plasmonic nanoparticle formulation was applied with 3 minutemassage, and the skin surface wiped with 3 applications of alternativewater and ethanol before laser irradiation. At 30 minutes followinglaser irradiation, no evidence of burning of the skin or erythema wasobserved in D or C, as Silica-coated particles could be sufficientlywiped from the skin surface. Magnified photography of D showed specificphotothermal damage (perifollicular erythema and edema) in thenanoparticle-targeted site.

FIG. 6 is a photograph showing nanoparticle-targeted photothermal damagein live human skin treated with a plasmonic nanoparticle formulation andclinical laser. A formulation of 1020 nm resonant, silica-coated (200nm-diameter) plasmonic nanoparticles in 20% propylene glycol and 3minute massage was contacted with live human skin. The procedure wasrepeated 3 times, and skin surface wiped with 3 applications ofalternating water and ethanol to remove residual particles. The treatedskin was irradiated with 1064 nm laser pulses (40 J/cm², 55 ms, 3passes). Following laser irradiation, clinical observation ofperifollicular erythema and edema was visible at hair follicles wherenanoparticles were targeted, but not visible in surrounding ornon-particle-treated tissues.

FIG. 7 is illustrative of a plasmonic nanoparticle formulation deliveryto human skin sebaceous gland. (A) Confocal microscope image of a humanskin biopsy and section, immunostained for Collagen IV basement membrane(blue) and PGP 9.5 nerve marker (green), shows hair follicle (HF) andsebaceous gland (SG) microanatomy. Red is silica nanoparticles (200 nm).(B) Schematic and dark field microscope image of excised human skintreated with plasmonic nanoparticle formulation, then sectioned andimaged horizontal to the follicle. Bright blue plasmonic particles arevisible up to 400 μm deep and within the human sebaceous gland.

FIG. 8 is illustrative of cosmetic formulations of plasmonicnanoparticles for sebaceous gland targeting that include surfactants.Silica-coated nanoparticles (200 nm diameter, 100 O.D.) were formulatedin 20% propylene glycol with the addition of surfactants sodium dodecylsulfate (SDS) or sodium laureth-2 sulfate (SLES), applied to human skinwith massage+ultrasound, and skin was sectioned in horizontal planes fordark field microscopy. (A) Formulations of plasmonic particles in 1%SDS/20% PG penetrated sebaceous gland down to 400 um as in FIG. 7. (B)Formulations of plasmonic particles in 1% SLES/20% PG penetratedsebaceous gland down to 600 um. Inset shows a skin section withoutvisible particles (scale bar 40 um). Sebaceous gland is pseudo-outlined.

FIG. 9 is an image depicting impact of massage vs. ultrasound onnanoparticle targeting to the human follicle and sebaceous gland.Silica-coated nanoparticles (200 nm diameter, 100 O.D.) were formulatedin 1% SDS/20% propylene glycol and applied to human skin with massage orultrasound. Dark field images of horizontal planar sections taken at low(20×) and high (50×) magnification show (A) little to no accumulation ofplasmonic particles into follicle infundibulum after massage alone,compared to (B) follicle infundibulum expansion and significantplasmonic particle accumulation after ultrasound alone.

FIG. 10 depicts an embodiment of the plasmonic nanoparticle cosmeticformulations for sebaceous gland targeting. Plasmonic nanoparticlescomprising different shapes and coatings were formulated in 1% SDS/20%propylene glycol and applied to human skin with massage+ultrasound, andskin was sectioned in horizontal planes for dark field microscopy. (A)Polyethylene glycol (PEG)-coated nanorods (gold, 15×30 nm dimension)were observed within the follicle infundibulum up to 200 um deep (whitearrow). (B) Lower concentration (10 O.D.) Silica-coated nanoplates(silver, 200 nm diameter) were observed up to 600 um deep in thefollicle and in the sebaceous gland (open arrow). Inset shows skinsections without visible particles (scale bar 100 um).

FIG. 11A is illustrative of temperature profiles of certain embodimentsof plasmonic nanoparticle formulations compared to other commercial andresearch chromophores.

FIG. 11B is illustrative of temperature profiles of certain embodimentsof plasmonic nanoparticle formulations compared to other commercial andresearch chromophores.

FIGS. 12A and 12B are images of embodiments of nanoparticle formulationsin porcine skin.

FIGS. 13A and 13B are images of biopsies taken from in vivo-treatedhuman skin, which were sectioned and immunostained for skin markers,with various embodiments of nanoparticles.

DETAILED DESCRIPTION

The biology of physiological and pathophysiological tissue growth andremodeling, and alterations in cell morphology is more complex thangenerally appreciated, involving an interacting network of biologicalcompounds, physical forces, and cell types.

An object of the subject matter described herein is to providecompositions, methods and systems for noninvasive and minimally-invasivetreatment of skin and underlying tissues, or other accessible tissuespaces with the use of nanoparticles. The treatment includes, but is notlimited to, hair removal, hair growth and regrowth, and skinrejuvenation or resurfacing, acne removal or reduction, wrinklereduction, pore reduction, ablation of cellulite and other dermal lipiddepositions, wart and fungus removal, thinning or removal of scarsincluding hypertrophic scars and keloids, abnormal pigmentation (such asport wine stains), tattoo removal, and skin inconsistencies (e.g. intexture, color, tone, elasticity, hydration). Other therapeutic orpreventative methods include but are not limited to treatment ofhyperhidrosis, anhidrosis, Frey's Syndrome (gustatory sweating), Homer'sSyndrome, and Ross Syndrome, actinici keratosis, keratosis follicularis,dermatitis, vitiligo, pityriasis, psoriasis, lichen planus, eczema,alopecia, psoriasis, malignant or non-malignant skin tumors.

Unless explained otherwise, all technical and scientific terms usedherein have the same meaning as commonly understood to one of ordinaryskill in the art to which this disclosure belongs. Although methods andmaterials similar or equivalent to those described herein can be used inthe practice or testing of the present disclosure, suitable methods andmaterials are described herein. The materials, methods, and examples areillustrative only and not intended to be limiting. Other features of thedisclosure are apparent from the following detailed description and theclaims.

“Administer” and “administration” as used herein, include providing orcausing the provision of a material to a subject, such as by a topical,subdermal, subcutaneous, intradermal, enteral, parenteral, rectal,nasal, intravenous, intramuscularly, intraperitoneal, or other route.

A “carrier suitable for administration” to a subject is any materialthat is physiologically compatible with a topical or route ofadministration to a desired vertebrate subject. Carriers can includesolid-based, dry materials for formulation; or the carrier can includeliquid or gel-based materials for formulations into liquid or gel forms.The specific type of carrier, as well as the final formulation depends,in part, upon the selected route(s) of administration and the type ofproduct.

A “comparable amount” is an amount that is measurably similar to a givenreference or standard.

The “components” of a formulation include any products or compoundsassociated with or contained within it.

An “effective dose”, “effective amount” or “therapeutic amount” is anamount sufficient to elicit the desired pharmacological, cosmetic ortherapeutic effects, thus resulting in effective prevention or treatmentof a disease or disorder, or providing a benefit in a vertebratesubject.

A “therapeutic effect” or “therapeutically desirable effect” refers to achange in a domain or region being treated such that it exhibits signsof being effected in the manner desired, e.g., cancer treatment causesthe destruction of tumor cells or halts the growth of tumor cells, acnetreatment causes a decrease in the number and/or severity of blemishes,hair removal treatment leads to evident hair loss, or wrinkle reductiontreatment causes wrinkles to disappear.

An “isolated” biological component (such as a nucleic acid molecule,protein, or cell) has been substantially separated or purified away fromother biological components in which the component was produced,including any other proteins, lipids, carbohydrates, and othercomponents.

A “nanoparticle”, as used herein, refers generally to a particle havingat least one of its dimensions from about 0.1 nm to about 9000 nm.

A “subject” or “patient” as used herein is any vertebrate species.

As used herein, a “substantially pure” or “substantially isolated”compound is substantially free of one or more other compounds.

A “target tissue” includes a region of an organism to which a physicalor chemical force or change is desired. As described herein, exemplarytarget tissues for acne treatment include a sebaceous gland, whileexemplary target tissues for hair removal include a pilosebaceous unit,a hair infundibulum, a hair follicle, or a non-follicular epidermis. A“region” of a target tissue includes one or more components of thetissue. Exemplary target tissue regions include the stem cell niche,bulge, sebaceous gland, dermal papilla, cortex, cuticle, inner rootsheath, outer root sheath, medulla, Huxley layer, Henle layer or pylorimuscle. A “domain” of a target tissue region includes basement membrane,extracellular matrix, cell-surface proteins, unbound proteins/analytes,glycomatrices, glycoproteins, or lipid bilayer.

A compound that is “substantially free” of some additional contents islargely or wholly without said contents.

A “plasmonic nanoparticle” is a nanometer-sized metallic structurewithin which localized surface plasmons are excited by light. Thesesurface plasmons are surface electromagnetic waves that propagate in adirection parallel to the metal/dielectric interface (e.g., metal/air ormetal/water).

A “light-absorbing nanomaterial” includes a nanomaterial capable ofdemonstrating a quantum size effect.

As described herein, provided are compositions that contain plasmonicnanoparticles to induce selective thermomodulation in a target tissue.

Plasmonic Nanoparticles.

Such compositions contain from about 2 to about 1×10¹⁸ nanoparticles(e.g., 10⁹ to about 10¹⁶ nanoparticles), such as 10², 10³, 10⁴, 10⁵,10⁶, 10⁷, 10⁸, 10⁹, 10¹⁰, 10¹¹, 10¹², 10¹³, 10¹⁴, 10¹⁵, 10¹⁶, 10¹⁷, or10¹⁸ particles. Preferably, the compositions contain about 10¹¹ to 10¹³particles so that the amount of particles localized to an effective 1 mltreatment volumes is from 10⁹ to 10¹¹. Generally, the compositionscontain nanoparticles in a concentration of from about 1 O.D. to about10,000 O.D. For embodiments wherein a greater concentration ofnanoparticles to a target region is desired, compositions containparticle concentrations with optical densities of, for example, 10O.D.-5000 O.D. more specifically 100 O.D.-1000 O.D., or opticaldensities greater than 1,000 O.D. In certain embodiments whereinincreased concentration of nanoparticles to a target region is desired,compositions contain particle concentrations with optical densities(O.D.) of 10 O.D.-1000 O.D., or optical densities greater than 1,000O.D. In some embodiments these correspond to concentrations of about1-10% w/w or more of nanoparticles. Determination of O.D. units in acomposition is determined using devices and analyses known in the art.

Nanoparticles may be homogenous or heterogeneous in size and othercharacteristics. The size of the nanoparticle is generally about 0.1 nmto about 50,000 nm (e.g., about 0.1 nm to about 5,000 nm) in at leastone dimension. Some variation in the size of a population ofnanoparticles is to be expected. For example, the variation might beless than 0.01%, 0.1%, 0.5%, 1%, 5%, 10%, 15%, 25%, 50%, 75%, 100%, 200%or greater than 200%. In certain embodiments where optimal plasmonicresonance is desired, a particle size in the range of from about 10 nmto about 100 nm is provided. Alternatively, in embodiments whereenhanced penetration of the nanoparticles into a target tissue regionsuch as a hair follicle is desired, a particle size in the range of fromabout 100 nm to about 1000 nm is provided. Modulation of particle sizepresent in the composition is also a useful means of concentrating thecomposition in a target domain. Further, as described herein,nanoparticles having a size range of from about 10 nm to about 100 nmcan be used as component of a larger molecular structure, generally inthe range of from about 100 nm to about 1000 nm. For example, theplasmonic nanoparticle can be surface coated to increase its size,embedded into an acceptable carrier, or it can be cross-linked oraggregated to other particles, or to other materials, that generate alarger particle. In certain embodiments where at least one dimension ofat least one nanoparticle within a solution of plasmonic nanoparticlesis below 50-100 nm, the nanoparticle surface can be coated with a matrix(e.g. silica) of 10-100 nm thickness or more in order to increase thatdimension or particle to 50-100 nm or more. This increased dimensionsize can increase the delivery of all nanoparticles to a target region(e.g., hair follicle) and limit delivery to non-target region (e.g.dermis). In one embodiment, the invention comprises a compositioncomprising at least about 1 O.D. (e.g., at least 10 O.D.) of coatedplasmonic nanoparticles (e.g., comprising silica or polyethylene glycol(PEG)) having a mean length in at least one dimension greater than about30 nanometers, wherein the coated nanoparticles are formulated in anacceptable carrier to be effective in induction of selectivethermoablation in a target tissue region with which the composition iscontacted, wherein the affinity of the coated nanoparticles for thetarget tissue region is substantially greater than the affinity of thecoated nanoparticles for a non-target tissue region.

Important considerations when generating nanoparticles include: 1) thezeta potential (positive, negative, or neutral) and charge density ofthe particles and resulting compositions; 2) thehydrophilicity/hydrophobicity of the particles and resultingcompositions; 3) the presence of an adsorption layer (e.g., a particleslippage plane); and 4) target cell adhesion properties. Nanoparticlesurfaces can be functionalized with thiolated moieties having negative,positive, or neutral charges (e.g. carboxylic acid, amine, hydroxyls) atvarious ratios. Moreover, anion-mediated surface coating (e.g. acrylate,citrate, and others), surfactant coating (e.g., sodium dodecyl sulfate,sodium laureth 2-sulfate, ammonium lauryl sulfate, sodiumoctech-1/deceth-1 sulfate, lecithin and other surfactants includingcetyl trimethylammonium bromide (CTAB), lipids, peptides), orprotein/peptide coatings (e.g. albumin, ovalbumin, egg protein, milkprotein, other food, plant, animal, bacteria, yeast, orrecombinantly-derived protein) can be employed. Block-copolymers arealso useful. Further, one will appreciate the utility of any othercompound or material that adheres to the surface of light-absorbingparticles to promote or deter specific molecular interactions andimprove particle entry into pores or follicles. In some embodiments, theparticle surface is unmodified. Modulation of hydrophilicity versushydrophobicity is performed by modifying nanoparticle surfaces withchemistries known in the art, including silanes, isothiocyanates, shortpolymers (e.g., PEG), or functionalized hydrocarbons. Polymer chains(e.g., biopolymers such as proteins, polysaccharides, lipids, andhybrids thereof; synthetic polymers such as polyethyleneglycol, PLGA,and others; and biopolymer-synthetic hybrids) of different lengths andpacking density are useful to vary the adsorption layer/slippage planeof particles.

Optical Absorption.

Preferred nanoparticles have optical absorption qualities of about 10 nmto about 10,000 nm, e.g., 100-500 nm, 500-750 nm, 600-900 nm, 700-1,000nm, 800-1,200 nm, or 500-2,000 nm. In specific embodiments, thenanoparticles have optical absorption useful to excitation by standardlaser devices or other light sources. For example, nanoparticles absorbat wavelengths of about 755 nm (alexandrite lasers), in the range ofabout 800-810 nm (diode lasers), or about 1064 nm (Nd: YAG lasers).Similarly, the nanoparticles absorb intense pulsed light (IPL), e.g., ata range of about 500 nm to about 1200 nm.

Assembly.

The nanoparticles provided herein can generally contain a collection ofunassembled nanoparticles. By “unassembled” nanoparticles it is meantthat nanoparticles in such a collection are not bound to each otherthrough a physical force or chemical bond either directly(particle-particle) or indirectly through some intermediary (e.g.particle-cell-particle, particle-protein-particle,particle-analyte-particle). In other embodiments, the nanoparticlecompositions are assembled into ordered arrays. In particular, suchordered arrays can include any three dimensional array. In someembodiments, only a portion of the nanoparticles are assembled, e.g., 5,10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 86, 90, 95,99% or greater than 99% of the nanoparticles are assembled in an orderedarray. The nanoparticles are assembled by a van der Walls attraction, aLondon force, a hydrogen bond, a dipole-dipole interaction, or acovalent bond, or a combination thereof.

“Ordered array” “Ordered arrays” can take the form of a macrostructurefrom individual parts that may be patterned or unpatterned in the formof spheres, colloids, beads, ovals, squares, rectangles, fibers, wires,rods, shells, thin films, or planar surface. In contrast, a “disorderedarray” lacks substantial macrostructure.

Geometrically Tuned Nanostructures.

The nanoparticles provided herein are formable in all shapes currentlyknown or to be created that absorb light and generate a plasmonresonance at a peak-wavelength or composition of wavelengths from 200 nmto 10,000 nm. In non-limiting examples, the nanoparticles are shaped asspheres, ovals, cylinders, squares, rectangles, rods, stars, tubes,pyramids, stars, prisms, triangles, branches, plates or comprised of aplanar surface. In non-limiting examples, the plasmonic particlescomprise nanoplates, solid nanoshells, hollow nanoshells nanorods,nanorice, nanospheres, nanofibers, nanowires, nanopyramids, nanoprisms,nanoplates or a combination thereof. Plasmonic particles present in thecomposition comprise a substantial amount of geometrically-tunednanostructures defined as 5, 10, 15, 25, 50, 75, 80, 85, 90, 95, 98, 99,99.9 or greater than 99.9% of particles.

Composition.

The nanoparticle is a metal (e.g., gold, silver), metallic composite(e.g., silver and silica, gold and silica), metal oxide (e.g. ironoxide, titanium oxide), metallic salt (e.g., potassium oxalate,strontium chloride), intermetallic (e.g., titanium aluminide, alnico),electric conductor (e.g., copper, aluminum), electric superconductor(e.g., yttrium barium copper oxide, bismuth strontium calcium copperoxide), electric semiconductor (e.g., silicon, germanium), dielectric(e.g., silica, plastic), or quantum dot (e.g., zinc sulfide, cadmiumselenium). In non-limiting examples, the materials are gold, silver,nickel, platinum, titanium, palladium, silicon, galadium. Alternatively,the nanoparticle contains a composite including a metal and adielectric, a metal and a semiconductor, or a metal, semiconductor anddielectric.

Coating.

Preferentially, the composition contains coated nanoparticles.

Type of Material Properties Exemplary Materials biorecognitive Moietywith affinity or Antibody, peptide, phage, material avidity for asubstrate DNA, RNA or analyte bioactive Moiety (e.g., protein, Growthfactor (e.g. VEGF), material analyte) that interrogates cytokine, cellsurface or modulates receptors, receptor ligands, the activity ofbiologic G-protein, kinase/ entity or cell phosphatase biologicalMaterial that is sourced albumin, ovalbumin, egg material from livingmatter protein, milk protein, other food, plant, animal, bacteria,yeast, or recombinantly- derived protein; peptides; enzymes, lipids,fatty acids, sugars biocide Material that is active in Synthetic ornatural material killing, destroying, or pesticides, synthetic ordisturbing biological natural anti-microbials matter dielectric Aninsulator that may be Silicon, doped materials polarized by an electricsemiconductors field chemo- Material that is able Receptor, receptorligand, recognitive to interactwith a moiety chemical molecule materialfor binding, biological or chemical reactions chemical Material thatcauses the Aldehyde, halogens, metals active material transformation ofa substance Polymer/ Long chain molecule PLGA, PEG, PEO, dendrimer(linear or branched, block polystyrene, carboxylate or co-block)styrene, rubbers, nylons, silicones, polysaccharides environmentallySurface molecule that Ph sensitive bond, light sensitive changes by itssensitive bond, heat sensitive polymer environment (e.g. acid) bond,enzyme sensitive bond, hydrolytic bond Hydrogel Polymer with highSynthetic 2-hydroxyethyl hydrophilicity and water metacrylate(HEMA)-based, “ordering” capacity polyethylene glycol (PEG)- based,PLGA, PEG- diacrylate; Natural ionic gels, alginate, gelatin, hyaluronicacids, fibrin Metal Thin metal coating to Gold, silver, nickel, achieveand/or improved platinum, titanium, and resonance palladium.functionalization capacity Semiconductors Semiconductor layer or Siliconand galadium. core that enhance Plasmon resonance polymer Fluorophorecross linked Fluorescein, rhodamine, containing to a polymer coat orCy5, Cy5.5, Cy7, Alexa a fluorescent directly to the surface dyes,Bodipy dyes marker of the particle Matrix Matrix coating that Silica,polyvinyl pyrrolidone, increases solubility of polysulfone,polyacrylamide, nanoparticles and/or polyethylene glycol, reduces“stickiness” polystyrene cellulose, to biological structures carbopol.

Biological Molecules.

The composition may contain a peptide, a nucleic acid, a protein, or anantibody. For example a protein, antibody, peptide, or nucleic acid thatbinds a protein of a follicular stem cell (e.g., keratin 15), a protein,glycomatrix, or lipid on the surface of a cell or stem cell, a protein,peptide, glycomatrix of the extracellular matrix or basement membrane.

Charged Moieties.

The coated nanoparticles may contain charged moieties whereby thosecharges mediate enhanced or diminished binding to components within oroutside the hair follicle via electrostatic or chemical interactions.

Class of Moiety Properties Exemplary Moieties Polar moieties Neutralcharge but increases Hydroxyl groups, hydrophilicity in waterisothiocyanates Non-polar moieties Increases hydrophobicity and orHydrocarbons, myristoylated improves solubility compounds, silanes,isothiocyanates Charged moieties Functional surface modificationsAmines, carboxylic acids, that change the zeta potential, hydroxylsisoelectric point, or pKa, and impact adsorption/binding tocomplementary charge compounds Ionic moieties Surface groups that have asingle Ammonium salts, chloride ion salts Basic moieties Groups thatdonate a hydrogen Amides, hydroxides, metal ions oxides, fluoride Acidicmoieties Moieties that accept hydrogen Carboxylic acids, sulfonic ionsacids, mineral acids Oxidative moieties Moieties that oxidize Manganeseions, reactive oxygen species Hydrophobic moieties Moieties that improvesolubility in Hydrocarbons, myristoylated non-aqueous solution and/orcompounds, silanes improve adsorption on the skin within a hair follicleHydrophilic moieties Moieties that are water-loving and PEG, PEO, PLGAprevent adsorption Agnostic moieties Moieties that bind a target cell,Antibodies, peptides, proteins structure, or protein of interestAntagonistic moieties Moieties that block the binding to Antibodies,peptides, proteins a target of interest Reactive moieties Moieties thatreact with biological Aldehydes or non-biological components with aresulting change in structure on the target

Description of Target Tissues.

Topical and Dermatological Applications.

Target tissues for topical and dermatological applications include thesurface of the skin, the epidermis and the dermis. Diseases orconditions suitable for treatment with topical and dermatologicalapplications include acne, warts, fungal infections, psoriasis, scarremoval, hair removal, hair growth, reduction of hypertrophic scars orkeloids, skin inconsistencies (e.g. texture, color, tone, elasticity,hydration), and malignant or non-malignant skin tumors.

As used herein, the term “acne” includes acne vulgaris as well as otherforms of acne and related cutaneous conditions, including acneaestivalis, acne conglobata, acne cosmetic, acne fulminans, acnekeloidalisnuchae, acne mechanica, acne miliarisnecrotica, acnenecrotica, chloracne, drug-induced acne, excoriated acne, halogen acne,lupus miliaris disseminates faciei, pomade acne, tar acne, and tropicalacne.

Subdermal Applications.

Target tissues for subdermal applications include the adipose tissue andconnective tissue below the integumentary system. Diseases or conditionssuitable for treatment with subdermatological applications includewrinkles and tattoos. Other applications include skin rejuvenationand/or resurfacing, the removal or reduction of stretch marks and fatablation.

Often, a specific region of the target tissue is a hair follicle, asebaceous gland, a merocrine sweat gland, an apocrine sweat gland, or anarrector pili muscle, within which a specific domain is targeted. Forexample, the bulge region of the hair follicle is targeted. Because inone embodiment the nanoparticles are useful to thermally ablate hairfollicle stem cells for hair removal, regions containing hair folliclestem cells are of particular interest for targeting. Thus, the targettissue region may include a stem cell niche, bulge, sebaceous gland,dermal papilla, cortex, cuticle, inner root sheath, outer root sheath,medulla, Huxley layer, Henle layer or pylori muscle. Each of theseregions may contain cells, stem cells, basement membrane, extracellularmatrix, growth factors, analytes, or other biologic components thatmediate hair follicle rejuvenation. Disruption or destruction of thesecomponents would have a therapeutic effect, e.g. slow or stop theprocesses that mediate hair regrowth, prevent the secretion of sebumfrom the sebaceous gland, damage or deter tumor cells, reduce theappearance of wrinkles. Structures can also be targeted that are inclose proximity to a desired target for ablation, especially whencapable of conducting heat effectively.

Localization Domains.

Provided are compositions containing nanoparticles that preferentiallylocalize to a domain of a target tissue region of a mammalian subject towhom the composition is administered.

Targeting Moieties.

The nanoparticles can be engineered to selectively bind to a domain ofthe target tissue. For example, the nanoparticles are operably linked tothe domain via a biologic moiety, in order to effectively target thenanoparticles to the target tissue domain. Preferably, the moietycontains a component of a stem cell, a progenitor cell, an extracellularmatrix component, a basement membrane component, a hair shaft component,a follicular epithelial component, or a non-follicular epidermalcomponent. Biological moieties include proteins such as cell surfacereceptors, glycoproteins or extracellular matrix proteins, as well ascarbohydrates, analytes, or nucleic acids (DNA, RNA) as well as membranecomponents (lipid bilayer components, microsomes).

Delocalization Domains.

Nanoparticles present in the composition preferentially delocalize awayfrom a domain of a target tissue region. Delocalization domains includespecific regions of a tissue into which nanoparticles do notsubstantially aggregate, or alternatively, are removed from the domainmore effectively. In preferred embodiments, the delocalization domain isa non-follicular epidermis, dermis, a component of a hair follicle(e.g., a hair stem cell, a stem cell niche, a bulge, a sebaceous gland,a dermal papilla, a cortex, a cuticle, an inner root sheath, an outerroot sheath, a medulla, a Huxley layer, a Henle layer, a pylori muscle),a hair follicle infundibulum, a sebaceous gland, a component of asebaceous gland, a sebocyte, a component of a sebocyte, or sebum

Energy Sources.

Provided herein are nonlinear excitation surface plasmon resonancesources, which include various light sources or optical sources.Exemplary light sources include a laser (ion laser, semiconductor laser,Q-switched laser, free-running laser, or fiber laser), light emittingdiode, lamp, the sun, a fluorescent light source or anelectroluminescent light source. Typically, the energy source is capableof emitting radiation at a wavelength from about 100, 200, 300, 400,500, 1000, 2000, 5000 nm to about 10,000 nm or more. The nonlinearexcitation surface plasmon resonance source is capable of emittingelectromagnetic radiation, ultrasound, thermal energy, electricalenergy, magnetic energy, or electrostatic energy. For example, theenergy is radiation at an intensity from about 0.00005 mW/cm² to about1000 TW/cm². The optimum intensity is chosen to induce high thermalgradients from plasmonic nanoparticles in regions from about 10 micronsto hundreds of microns in the surrounding tissue, but has minimalresidual effect on heating tissue in which particles do not residewithin a radius of about 100 microns or more from the nanoparticle. Incertain embodiments, a differential heat gradient between the targettissue region and other tissue regions (e.g., the skin) is greater than2-fold, 3-fold, 5-fold, 10-fold, 15-fold, 20-fold, 50-fold, 100-fold, orgreater than 100 fold.

The energy can be tuned by monitoring thermal heat gradients on thesurface of the skin with a thermal/infrared camera. As demonstratedherein, the methods and systems of the present disclosure providesuperior efficacy when a surface plasmon is generated on thenanoparticles by the action of the radiation. Typically, the plasmon isgenerated in a one-photon mode or, alternatively, a two-photon mode, amulti-photon mode, a step-wise mode, or an up-conversion mode.

Delivery of Radiation.

Physical means of delivery of the energy from the nonlinear excitationsurface plasmon resonance source to the target tissue region include afiber, waveguide, a contact tip or a combination thereof.

Optical sources include a CW optical source or a pulsed optical source,which may be a single wavelength polarized (or, alternatively,unpolarized) optical source capable of emitting radiation at a frequencyfrom about 200 nm to about 10,000 nm. Alternatively, the optical sourceis a multiple wavelength polarized (or, alternatively, unpolarized)optical source capable of emitting radiation at a wavelength from about200 nm to about 10,000 nm. The pulsed optical source is generallycapable of emitting pulsed radiation at a frequency from about 1 Hz toabout 1 THz. The pulsed optical source is capable of a pulse less than amillisecond, microsecond, nanosecond, picoseconds, or femtosecond induration. For example, a source emitting radiation at a wavelength of755 nm is operated in pulse mode such that the emitted radiation ispulsed at a duration of 0.25-300 milliseconds (ms) per pulse, with apulse frequency of 1-10 Hz. In another example, radiation emitted at awavelength of 810 nm is pulsed at 5-100 ms with a frequency of 1-10 Hz.In a further example, a source emitting radiation at a wavelength of1064 nm is pulsed at 0.25-300 ms at a frequency of 1-10 Hz. In yetanother example, a source emitting intense pulsed light at a wavelengthof 530-1200 nm is pulsed at 0.5-300 ms at a frequency of 1-10 Hz. Theoptical source may be coupled to a skin surface cooling device to reduceheating of particles or structures on the skin surface and focus heatingto components within follicles or tissue structures at deeper layers.

Nanoparticle-Containing Compositions.

In order to provide optimal dermal penetration into the target tissue,the plasmonic nanoparticles in certain embodiments are formulated invarious compositions. Preferentially, the nanoparticles are formulatedin compositions containing 1-10% v/v surfactants (e.g. sodium dodecylsulfate, sodium laureth 2-sulfate, ammonium lauryl sulfate, sodiumoctech-1/deceth-1 sulfate). Surfactants disrupt and emulsify sebum orother hydrophobic fluids to enable improved targeting of hydrophilicnanoparticles to the hair follicle, infundibulum, sebaceous gland, orother regions of the skin. Surfactants also lower the free energynecessary to deliver hydrophilic nanoparticles into small hydrophobiccrevices such as the space between the hair shaft and follicle or intothe sebaceous gland. Nanoparticle-containing compositions may alsoinclude emulsions at various concentrations (1-20% w/v) in aqueoussolutions, silicone/oil solvents, polypropylene gel, propylene glycol orcreams (e.g. comprising alcohols, oils, paraffins, colloidal silicas).In other embodiments, the formulation contains a degradable ornon-degradable polymer, e.g., synthetic polylactide/co-glycolideco-polymer, porous lauryllactame/caprolactame nylon co-polymer,hydroxyethylcellulose, polyelectrolyte monolayers, or alternatively, innatural hydrogels such as hyaluronic acid, gelatin and others. Infurther embodiments, a hydrogel PLGA, PEG-acrylate is included in theformulation. Alternatively, a matrix component such as silica,polystyrene or polyethylene glycol is provided in the formulation. Otherformulations include components of surfactants, a lipid bilayer, aliposome, or a microsome. A nanoparticle may comprise a largermicron-sized particle.

Effective Doses.

As described herein, an effective dose of the nanoparticle-containingcompositions includes an amount of particles required, in some aspects,to generate an effective heat gradient in a target tissue region, suchthat a portion of the target tissue region is acted upon by thermalenergy from excited nanoparticles. A “minimal effective dose” is thesmallest number or lowest concentration of nanoparticles in acomposition that are effective to achieve the desired biological,physical and/or therapeutic effect(s). Preferentially, the plasmonicnanoparticles have an optical density of 10 O.D.-1,000 O.D. at one or aplurality of peak resonance wavelengths.

Cosmetically Acceptable Carriers.

Provided are cosmetic or pharmaceutical compositions with a plurality ofplasmonic nanoparticles and a cosmetically or pharmaceuticallyacceptable carrier. Generally, the carrier and composition must besuitable for topical administration to the skin of a mammalian subject,such that the plasmonic nanoparticles are present in an effective amountfor selective thermomodulation of a component of the skin.Preferentially, the nanoparticles are formulated with a carriercontaining 1-10% v/v surfactants (e.g. sodium dodecyl sulfate, sodiumlaureth 2-sulfate, ammonium lauryl sulfate, sodium octech-1/deceth-1sulfate) to enable disruption of the epidermal skin barrier, emulsifysebum, improve mixing of hydrophilic nanoparticles with hydrophobicsolutions, and reduce entropic barriers to delivering hydrophilicparticles to hydrophobic regions of the skin (e.g. between the hairshaft and surrounding sheath or follicle). In some embodiments, thecarrier contains a polar or non-polar solvent. For example, suitablesolvents include alcohols (e.g., n-Butanol, isopropanol, n-Propanol,Ethanol, Methanol), hydrocarbons (e.g., pentane, cyclopentane, hexane,cyclohexane, benzene, toluene, 1,4-Dioxane), chloroform, Diethyl-ether,water, water with propylene glycol, acids (e.g., acetic acid, formicacid), bases, acetone, isooctanes, dimethyl sulfoxide (DMSO),dimethylformamide (DMF), acetonitrile (MeCN), tetrahydrofuran (THF),dichloromethane (DCM), ethylacetate, tetramethylammonium hydroxide,isopropanol, and others. In other embodiments, a stabilizing agent suchas antioxidants, preventing unwanted oxidation of materials,sequestrants, forming chelate complexes and inactivating traces of metalions that would otherwise act as catalysts, emulsifiers, ionic ornon-ionic surfactants, cholesterol or phospholipids, for stabilizationof emulsions (e.g. egg yolk lecithin, Sodium stearoyllactylate, sodiumbis(2-ethylhexyl-sulfosuccinate (AOT)), ultraviolet stabilizers,protecting materials, especially plastics, from harmful effects ofultraviolet radiation is provided. In further embodiments, a compositionwith a cosmetically acceptable carrier is generated such that thenanoparticles are substantially in a suspension.

Other components are also optionally included, including an emulsion,polymer, hydrogel, matrix, lipid bilayer, liposome, or microsome.Additionally, inclusion of a detectable colorant (e.g., a pigment), afragrance, a moisturizer, and/or a skin protectant is optional. In someexamples, the formulation has a viscosity of above, below or within0.1-10,000 (e.g., 5e⁻⁴×10³, 1,000), as measured in millipascal-seconds(mPa·s).

Nanoparticle quantities per milliliter in a composition are subject tomodification for specific binding and can range from 10⁹ to 10¹⁸particles but generally about 10¹¹ to 10¹³ nanoparticles per milliliter.Nanoparticle quantities per milliliter in a formulation are subject tomodification for specific binding but generally up to about 10²³nanoparticles per milliliter. In certain embodiments wherein increasedconcentration of nanoparticles to a target region is desired,compositions contain particle concentrations with optical densities of10 O.D.-1000 O.D., or optical densities greater than 1,000 O.D. In someembodiments these correspond to concentrations of about 0.1-10% w/w ormore of nanoparticles.

Prior to application of nanoparticle formulations, skin and hairfollicles can be pre-treated to increase the delivery of nanoparticlesto a target region. In some embodiments, hair shafts are cut or removedvia shaving, waxing, cyanoacrylate surface peels, calcium thioglycolatetreatment, or other techniques to remove the hair shaft and/or hairfollicle plugs and create a void wherein nanoparticles can accumulate.Orifices of active or inactive follicles can be blocked by plugs formedof corneocytes and/or other material (e.g. cell debris, soot,hydrocarbons, cosmetics). In some embodiments pre-treatment with surfaceexfoliation including mechanical exfoliation (e.g., salt glow ormicrodermabrasion) and chemical exfoliation (e.g., enzymes, alphahydroxyacids, or betahydroxy acids) removes plugs from the orifice of folliclesto increase the targeting of nanoparticle formulations to target regionswithin the hair follicle.

In some embodiments, the nanoparticle formulations are formulated forapplication by a sponge applicator, cloth applicator, direct contact viaa hand or gloved hand, spray, aerosol, vacuum suction, high pressure airflow, or high pressure liquid flow, roller, brush, planar surface,semi-planar surface, wax, ultrasound and other sonic forces, mechanicalvibrations, physical manipulation, hair shaft manipulation (includingpulling, massaging), physical force, electrophoresis, iontophoresis,thermal manipulation, and other treatments. In some embodiments,nanoparticle formulation treatments are performed alone, in combination,sequentially or repeated 1-24 times. In other embodiments, the plasmonicnanoparticles are capable of selectively localizing to a first componentof the skin, where physical massage or pressure, ultrasound, or heatincrease the selective localization of the nanoparticles to this firstcomponent. Additionally, the nanoparticles are selectively removablefrom components of the skin other than the first component, such removalaccomplished with acetone, alcohol, water, air, peeling of the skin,chemical peeling, waxing, or reduction of the plasmonic compound.Further, in some embodiments the nanoparticles have a coat layer toincrease solubility of the nanoparticles in the carrier and/or reduce“stickiness” and accumulation in non-target areas. The subject matterdescribed herein also provides embodiments in which at least a portionof an exterior surface of the nanoparticle is modified, such as toinclude a layer of a polymer, polar monomer, non-polar monomer, biologiccompound, a metal (e.g., metallic thin film, metallic composite, metaloxide, or metallic salt), a dielectric, or a semiconductor.Alternatively, the exterior surface modification is polar, non-polar,charged, ionic, basic, acidic, reactive, hydrophobic, hydrophilic,agonistic, or antagonistic. In certain embodiments where at least onedimension of at least one nanoparticle within a solution of plasmonicnanoparticles is below 50-100 nm, the nanoparticle surface can be coatedwith a matrix (e.g. silica) of 10-100 nm thickness or more in order toincrease that dimension or particle to 50-100 nm or more. This increaseddimension size can increase the delivery of all nanoparticles to atarget region (e.g., hair follicle) and limit delivery to non-targetregion (e.g. dermis).

Penetration Means.

Preferably, the compositions of the instant disclosure are topicallyadministered. Provided herein area means to redistribute plasmonicparticles from the skin surface to a component of dermal tissueincluding a hair follicle, a component of a hair follicle, a follicleinfundibulum, a sebaceous gland, or a component of a sebaceous glandusing high frequency ultrasound, low frequency ultrasound, massage,iontophoresis, high pressure air flow, high pressure liquid flow,vacuum, pre-treatment with Fractionated Photothermolysis laser orderm-abrasion, or a combination thereof. The nanoparticles describedherein are formulated to penetrate much deeper—up to severalcentimeters, or into the panniculus adiposus (hypodermis) layer ofsubcutaneous tissue. For example, the compositions can be administeredby use of a sponge applicator, cloth applicator, spray, aerosol, vacuumsuction, high pressure air flow, high pressure liquid flow directcontact by hand ultrasound and other sonic forces, mechanicalvibrations, physical manipulation, hair shaft manipulation (includingpulling, massaging), physical force, thermal manipulation, or othertreatments. Nanoparticle formulation treatments are performed alone, incombination, sequentially or repeated 1-24 times.

Cosmetic and Therapeutic Uses of Plasmonic Nanoparticles.

In general terms, Applicant(s) have created systems and methods for thecosmetic and therapeutic treatment of dermatological conditions,diseases and disorders using nanoparticle-based treatments methods.

Acne Treatment.

Acne is caused by a combination of diet, hormonal imbalance, bacterialinfection (Propionibacterium acnes), genetic predisposition, and otherfactors. The nanoparticle-based methods and systems described herein foracne treatment are able to focally target causative regions of thedermis, the sebaceous gland and the hair follicle, and thus haveadvantages compared to the existing techniques known in the art,including chemical treatment (peroxides, hormones, antibiotics,retinoids, and anti-inflammatory compounds), dermabrasion, phototherapy(lasers, blue and red light treatment, or photodynamic treatment), orsurgical procedures.

In particular, laser-based techniques are becoming an increasinglypopular acne treatment, but a substantial limitation is the lack ofselective absorptive properties among natural pigments (e.g. fat, sebum)for specific wavelengths of light such that selective heating of onecell, structure, or component of tissue, particularly in the sebaceousglands, infundibulum, and regions of the hair follicle, is not achievedwithout heating of adjacent off-target tissue. The nanoparticlesdescribed herein provide significantly higher photothermal conversionthan natural pigments enabling laser energy to be focused to specificcells, structures, or components of tissue within the sebaceous gland,infundibulum, or regions of the hair follicle for selective photothermaldamage.

Using the materials and techniques described herein may provide acnetreatments of greater duration than existing methodologies. In certainembodiments, tuned selective ablation of the sebaceous gland orinfundibulum is achieved as described herein. In particular, plasmonicnanoparticles are specifically localized to regions of hair follicles inor proximate to the sebaceous gland or infundibulum.

Plasmonic nanoparticles exhibit strong absorption at wavelengths emittedby standard laser hair removal devices (e.g., 755 nm, 810 nm, 1064 nm)relative to surrounding epidermal tissue. Thus, irradiation of targetedplasmonic nanoparticles with laser light induces heat radiation from theparticles to the adjacent sebum, sebaceous gland, infundibulum, andother acne causing agents.

Hair Removal.

The nanoparticle-based methods and systems described herein for skintreatment have advantages compared to the existing techniques known inthe art, including laser-based techniques, chemical techniques,electrolysis, electromagnetic wave techniques, and mechanical techniques(e.g., waxing, tweezers). Such techniques fail to adequately providepermanent hair removal across a breadth of subjects. In particular,subjects having light to medium-pigmented hair are not adequately servedby these techniques, which suffer from side-effects including pain andthe lack of beneficial cosmetic affects including hair removal.Laser-based techniques are popular in a variety of applications, but asubstantial limitation is the lack of selective absorptive propertiesamong natural pigments (e.g. melanin) for specific wavelengths of lightsuch that selective heating of one cell, structure, or component oftissue is achieved without heating of adjacent off-target tissues. Thenanoparticles described herein provide significantly higher photothermalconversion than natural pigments enabling laser energy to be focused tospecific cells, structures, or components of tissue for selectivephotothermal damage. The methods described herein are useful for hairremoval of all types and pigmentations. For example, melanin, thepredominant hair pigment, is an aggregation of chemical moietiesincluding eumelanin and phaeomelanin. Eumelanin colors hair grey, black,yellow, and brown. A small amount of black eumelanin in the absence ofother pigments causes grey hair. Types of eumelanin include blackeumelanin and brown eumelanin, with black melanin being darker thanbrown. Generally, black eumelanin predominates in non-European subjectsand aged Europeans, while brown eumelanin is in greater abundance inyoung European subjects. Phaeomelanin predominates in red hair. Inanother example, vellus hair (“peach fuzz”) is a type of short, fine,light-colored, and usually barely noticeable hair that develops on muchor most of a subject's body (excluding lips, palms of hand, sole offoot, navel and scar tissue). While the density of vellus hair isgenerally lower than that of other hair types, there is variation fromperson to person in the density, thickness, and pigmentation. Vellushair is usually less than 2 mm long and the follicle containing thevellus hair is generally not connected to a sebaceous gland. Conditionsassociated with an overabundance of vellus hair include Cushing'ssyndrome and anorexia nervosa, such overgrowth being treatable using themethods and compositions described herein. Further, provided are methodsof targeting hair growth at a given stage. Hair grows in cycles ofvarious stages or phases. Growth phase is termed “anagen”, while“catagen” includes the involuting or regressing phase, and “telogen”encompasses the resting or quiescent phase. Each phase has severalmorphologically and histologically distinguishable sub-phases.Generally, up to 90% of the hair follicles on a subject are in anagenphase (10-14% are in telogen and 1-2% in catagen). The cycle's length isgoverned by cytokines and hormones, and varies on different parts of thebody. For eyebrows, the cycle is completed in around 4 months, while ittakes the scalp 3-4 years to finish. The methods and compositionsdescribed herein are sufficient to treat hair of all growth stages orphases.

More permanent reduction or removal of all hair types is providedherein, relative to hair removal treatments known in the art. In certainembodiments, tuned selective ablation of the hair shaft and destructionof stem cells in the bulge region is provided, as described herein. Inparticular, plasmonic nanoparticles are specifically localized toregions of hair follicles in or proximate to the bulge region, a stemcell-rich domain of the hair follicle. Moreover, the plasmonicnanoparticles are localized in close approximation of ˜50-75% of thehair shaft structure.

Plasmonic nanoparticles exhibit strong absorption at wavelengths emittedby standard laser hair removal devices (e.g., 755 nm, 810 nm, 1064 nm)relative to surrounding epidermal tissue. Thus, irradiation of targetedplasmonic nanoparticles with laser light induces heat radiation from theparticles to the adjacent stem cells (or in some cases, the architectureof the hair shaft itself), resulting in cell death and a disruption ofthe normal regenerative pathway.

Non-Malignant and Malignant Skin Tumors.

Laser therapies for the prevention and treatment of non-malignant,malignant, melanoma and non-melanoma skin cancers have been focusedlargely on photodynamic therapy approaches, whereby photosensitiveporphyrins are applied to skin and used to localize laser light, producereactive oxygen species and destroy cancer cells via toxic radicals. Forexample, 5-ALA combined with laser treatment has been FDA-approved forthe treatment of non-melanoma skin cancer actinic keratoses, and it isused off-label for the treatment of widely disseminated, surgicallyuntreatable, or recurrent basal cell carcinomas (BCC). However, thisprocedure causes patients to experiences photosensitivity, burning,peeling, scarring, hypo- and hyper-pigmentation and other side effectsdue to non-specific transdermal uptake of porphyrin molecules. Thenanoparticles described herein provide significantly higher photothermalconversion than natural pigments and dyes, enabling laser energy to befocused to specific cells, structures, or components of tissue forselective thermomodulation

Using the materials and techniques described herein may provide cancertreatments of greater degree and duration than existing methodologies.In certain embodiments, tuned selective ablation of specific targetcells, such as Merkel cells or Langerhans cells, as described herein. Inparticular, plasmonic nanoparticles are specifically localized toregions of hair follicles where follicular bulge stem cells arise toform nodular basal cell carcinomas and other carcinomas. Plasmonicnanoparticles may also be delivered to other target cells that causetumors, for example, the interfollicular epithelium, which include thecell of origin for superficial basal cell carcinomas.

Plasmonic nanoparticles exhibit strong absorption at wavelengths emittedby standard laser hair removal devices (e.g., 755 nm, 810 nm, 1064 nm)relative to surrounding epidermal tissue. Thus, irradiation of targetedplasmonic nanoparticles with laser light induces heat radiation from theparticles to the adjacent keratinocyte, melanocyte, follicular bulgestem cell, cancer cell, or cancer cell precursor, resulting in celldeath or inhibited cell growth for cancer prevention and treatment.

Subdermal Applications.

Target tissues for subdermal applications include the adipose tissue andconnective tissue below the integumentary system. Diseases or conditionssuitable for treatment with subdermatological applications includewrinkles and tattoos. Other applications include skin rejuvenationand/or resurfacing, the removal or reduction of stretch marks and fatablation.

Vascular Applications.

Target tissues for vascular applications include arteries, arterioles,capillaries, vascular endothelial cells, vascular smooth muscle cells,veins, and venules. Diseases or conditions suitable for treatment withvascular applications include spider veins, leaky valves, and vascularstenosis. In particular, vein abnormalities account for a substantialproportion of cosmetic diseases or conditions affecting the vasculature.Individuals with vein abnormalities such as spider veins or faultyvenous valves suffer from pain, itchiness, or undesirable aesthetics.

Additionally, there are several indication for which ablation of othervessels including arteries, arterioles, or capillaries could providetherapeutic or cosmetic benefit including: 1) ablation of vasculaturesupplying fat pads and/or fat cells, 2) ablation of vasculaturesupporting tumors/cancer cells, 3) ablation of vascular birth marks(port-wine stains, hemangiomas, macular stains), and 4) any otherindication whereby ablation of vessels mediates the destruction oftissue and apoptosis or necrosis of cells supported by those vesselswith therapeutic or cosmetic benefit. Provided herein are methods forusing the compositions described herein for the selective destruction ofcomponent(s) of veins from plasmonic nanoparticles focally or diffuselydistributed in the blood. Plasmonic nanoparticles are combined with apharmaceutically acceptable carrier as described above and areintroduced into the body via intravenous injection. Nanoparticlesdiffuse into the blood and, in some embodiments, localize to specificvascular tissues. Subsequently, the nanoparticles are activated withlaser or light-based systems as known in the art for treating skinconditions such as hair removal or spider vein ablation. Alternatively,image or non-image guided fiber optic waveguide-based laser or lightsystems may be used to ablate vessel or blood components in largerveins. In one embodiment, a device with dual functions for bothinjecting nanoparticles and administering light through on opticalwaveguide may be used. Activated nanoparticles heat blood and adjacenttissue (vessels, vessel walls, endothelial cells, components on or inendothelial cells, components comprising endothelial basement membrane,supporting mesenchymal tissues, cells, or cell components around thevessel, blood cells, blood cell components, other blood components) toablative temperatures (38-50 degrees C. or higher).

Provided herein is a composition comprising a pharmaceuticallyacceptable carrier and a plurality of plasmonic nanoparticles in anamount effective to induce thermomodulation of a vascular orintravascular target tissue region with which the composition isintravenously contacted. Furthermore, the composition of plasmonicnanoparticle may comprise a microvascular targeting means selected fromthe group consisting of anti-microvascular endothelial cell antibodiesand ligands for microvascular endothelial cell surface receptors. Alsoprovided is a method for performing thermoablation of a target vasculartissue in a mammalian subject, comprising the steps of contacting aregion of the target vascular tissue with a composition comprising aplurality of plasmonic nanoparticles and a pharmaceutically acceptablecarrier under conditions such that an effective amount of the plasmonicnanoparticles localize to a domain of the target vascular region; andexposing the target tissue region to energy delivered from a nonlinearexcitation surface plasmon resonance source in an amount effective toinduce thermoablation of the domain of the target vascular region.

Oral and Nasal Applications.

Target tissues for oral applications include the mouth, nose, pharynx,larynx, and trachea. Diseases or conditions suitable for treatment withvascular applications include oral cancer, polyps, throat cancer, nasalcancer, and Mounier-Kuhn syndrome.

Endoscopic Applications.

Target tissues for endoscopic applications include the stomach, smallintestine, large intestine, rectum and anus. Diseases or conditionssuitable for treatment with vascular applications includegastrointestinal cancer, ulcerative colitis, Crohn's disease, IrritableBowel Syndrome, Celiac Disease, Short Bowel Sydrome, or an infectiousdisease such as giardiasis, tropical sprue, tapeworm infection,ascariasis, enteritis, ulcers, Whipple's disease, and megacolon.

Methods of Thermomodulation.

Provided are methods for performing thermomodulation of a target tissueregion. A nanoparticle composition comprising a plurality of plasmonicnanoparticles under conditions such that an effective amount of theplasmonic nanoparticles localize to a domain of the target tissueregion; and exposing the target tissue region to energy delivered from anonlinear excitation surface plasmon resonance source in an amounteffective to induce thermomodulation of the domain of the target tissueregion.

Removal of Non-Specifically Bound Nanoparticles.

Removing nanoparticles localized on the surface of the skin may beperformed by contacting the skin with acetone, alcohol, water, air, adebriding agent, or wax. Alternatively, physical debridement may beperformed. Alternatively, one can perform a reduction of the plasmoniccompound.

Amount of Energy Provided.

Skin is irradiated at a fluence of 1-60 Joules per cm² with laserwavelengths of about, e.g., 750 nm, 810 nm, 1064 nm, or otherwavelengths, particularly in the range of infrared light. Variousrepetition rates are used from continuous to pulsed, e.g., at 1-10 Hz,10-100 Hz, 100-1000 Hz. While some energy is reflected, it is anadvantage of the subject matter described herein is that a substantialamount of energy is absorbed by particles, with a lesser amount absorbedby skin. Nanoparticles are delivered to the hair follicle, infundibulum,or sebaceous gland at concentration sufficient to absorb, e.g., 1.1-100×more energy than other components of the skin of similar volume. This isachieved in some embodiments by having a concentration of particles inthe hair follicle with absorbance at the laser peak of 1.1-100× relativeto other skin components of similar volume.

To enable tunable destruction of target skin structures (e.g., sebaceousglands, infundibulum, hair follicles), light-absorbing nanoparticles areutilized in conjunction with a laser or other excitation source of theappropriate wavelength. The laser light may be applied continuously orin pulses with a single or multiple pulses of light. The intensity ofheating and distance over which photothermal damage will occur arecontrolled by the intensity and duration of light exposure. In someembodiments, pulsed lasers are utilized in order to provide localizedthermal destruction. In some such embodiments, pulses of varyingdurations are provided to localize thermal damage regions to within0.05, 0.1, 0.5, 1, 2, 5, 10, 20, 30, 50, 75, 100, 200, 300, 500, 1000microns of the particles. Pulses are at least femtoseconds, picoseconds,microseconds, or milliseconds in duration. In some embodiments, the peaktemperature realized in tissue from nanoparticle heating is at least 5,10, 15, 20, 25, 30, 40, 50, 60, 70, 80, 90, 100, 200, 300, or 500degrees Celsius. In some embodiments that utilize pulsed heating, highpeak temperatures are realized locally within the hair shaft withoutraising the macroscopic tissue temperature more than 0.1, 0.5, 1, 2, 3,4, 5, 7, 9, 12, 15, or 20 degrees Celsius. In some embodiments shortpulses (100 nanoseconds-1000 microseconds) are used to drive very hightransient heat gradients in and around the target skin structure (e.g.,sebaceous gland and/or hair follicle) from embedded particles tolocalize damage in close proximity to particle location. In otherembodiments, longer pulse lengths (1-10 ms, or 1-500 ms) are used todrive heat gradients further from the target structure to localizethermal energy to stem cells in the bulge region or other componentsgreater than 100 μm away from the localized particles. Fluences of 1-10Joules per cm² or 1-30 Joules per cm² are generally sufficient tothermally ablate follicles that have high particle concentrations andthus higher absorbance than skin (e.g., 1.1-100 times per volumeabsorbance of skin). These fluences are often lower than what iscurrently employed (e.g., Diode: 25-40 J/cm², Alexandrite: 20 J/cm2,Nd:YAG: 30-60 J/cm²) and lead to less damage to non-follicular regions,and potentially less pain.

Plasmon Resonance Systems.

Provided are plasmon resonance systems containing a surface thatincludes a plurality of plasmonic nanoparticles, and a nonlinearexcitation source. Optionally, the system contains a means to generatethermal heating of the surface. Preferably, the surface is a componentof skin that is targeted for cosmetic or therapeutic treatment (e.g.,bulge region for hair removal, infundibulum or sebaceous gland for acneprevention). Also provided as a component of the system is a means fordelivering plasmonic nanoparticles to the skin surface, such as anapplicator, a spray, an aerosol, vacuum suction, high pressure air flow,or high pressure liquid flow. Further provided are means of localizingplasmonic nanoparticles to a component of the skin (e.g., hair follicle,bulge region, sebaceous gland, infundibulum). Useful surface deliverymeans include a device that generates high frequency ultrasound, lowfrequency ultrasound, heat, massage, contact pressure, or a combinationthereof.

Further provided are systems that contain a removal means for removingnanoparticles on a non-follicular portion of the skin. The removal meansincludes at least one of acetone, alcohol, water, air, chemical peeling,wax, or a compound that reduces the plasmonic compound.

In addition, the systems of the present disclosure provide nonlinearexcitation source that generates a continuous wave optical source or apulsed optical source. Alternatively, the nonlinear excitation source iscapable of generating electromagnetic radiation, ultrasound, thermalenergy, electrical energy, magnetic energy, or electrostatic energy.Provided are systems wherein the nonlinear excitation source is capableof irradiating the nanoparticles with an intensity from about 0.00005mW/cm² to about 1000 TW/cm². Further, the nonlinear excitation source iscapable of functioning in a one-photon mode, two-photon mode,multi-photon mode, step-wise mode, or up-conversion mode. A fiber, awaveguide, a contact tip, or a combination thereof may be used in theinstant systems.

In some embodiments, the system contains a monitoring device such as atemperature sensor or a thermal energy detector. In other embodiments,the systems also contain a controller means for modulating the nonlinearexcitation source (e.g., a “feedback loop controller”). In a relatedembodiment, the system contains a means for detecting a temperature ofthe surface or a target tissue adjacent to the surface, wherein thecontroller means modulates the intensity of the nonlinear excitationsource and/or the duration of the excitation. In such embodiments, thecontroller means preferably modulates the intensity of the nonlinearexcitation source such that a first component of the hair follicle isselectively thermoablated relative to a second component of the hairfollicle. In further embodiments, a cooling device is directly contactedwith the skin during irradiation to minimize the heating ofnanoparticles or skin at the surface, while nanoparticles that havepenetrate more deeply into the follicle, skin, or sebaceous gland heatto temperatures that selectively ablate the adjacent tissues.

Skin is an exemplary target tissue. The skin preferably contains a hairfollicle and/or a sebaceous gland, where the nonlinear excitation sourcegenerates energy that results in heating the skin in an amount effectiveto induce thermomodulation of a hair follicle, a infundibulum, asebaceous gland, or a component thereof, such as by heating sufficientto cause the temperature of the skin to exceed 37° C., such as 38° C.,39° C., 40° C., 41° C., 42° C., 43° C., 44° C., 45° C., 46° C., 47° C.,48° C., 49° C., to about 50° C. or greater.

Methods of Formulation.

Also provided are methods for formulating the nanoparticles of thepresent disclosure into a form suitable for use as described herein. Inparticular, the nanoparticle compositions are generated by:

-   -   a) forming a first mixture containing a plurality of        nanoparticles and a first solvent;    -   b) exchanging the first solvent for a second solvent to form a        second mixture; and    -   c) combining the second mixture and a cosmetically or        pharmaceutically acceptable carrier; thereby forming a        nanoparticle composition.

The exchanging step is optionally performed using liquid chromatography,a solvent exchange system, a centrifuge, precipitation, or dialysis.Preferably, the nanoparticles are surface modified through a controlledreduction step or an oxidation step. Such surface modification mayinvolve a coating step, such as the adsorbance of a monomer, polymer, orbiological entity to a surface of the nanoparticle. Typically, thecoating step involves contacting the nanoparticles with an oxidativeenvironment. Further, the coating step may include monomerpolymerization to create polymer coat.

The methods described herein may also include the steps of dissolvingthe nanoparticles in a non-polar solvent and subsequently mixing thedissolved nanoparticles with a polar solvent so as to encapsulate thenanoparticles in an emulsion. Further, the addition of surfactants (e.g.sodium dodecyl sulfate, sodium laureth 2-sulfate, ammonium laurylsulfate, sodium octech-1/deceth-1 sulfate) at concentrations of 0.1-10%may be used to disrupt the epidermal skin barrier, emulsify the sebumand enable improved mixing of hydrophilic nanoparticles in aqueoussolutions. Further, a concentration of the nanoparticles such ascentrifugation or lyophilization may be employed. Further, thenanoparticles may be pretreated with heat or radiation. Also provided isthe optional step of conjugating a biological entity or plurality ofbiological entities to the nanoparticles. Such a conjugating step mayinvolve a thiol, amine, or carboxyl linkage of the biological entitiesto the nanoparticles.

Diseases and Disorders.

The present disclosure can be used on human (or other animal) skin forthe treatment of wrinkles and other changes related to photo-aging orchronologic aging (generally termed skin rejuvenation), for thetreatment of diseases including skin diseases, for the reduction of acneand related disorders such as rosacea, folliculitis, pseudofolliculitisbarbae or proliferative or papulosquamous disorders such as psoriasis,for the stimulation or reduction of hair growth, and for reduction ofcellulite, warts, hypopigmentation such as port-wine stain (PWS; nevusflammeus), birthmarks, hyperhidrosis, varicose veins, pigment problems,tattoos, vitiligo, melasma, scars, stretch marks, fungal infections,bacterial infections, dermatological inflammatory disorders,musculoskeletal problems (for example, tendonitis or arthritis), toimprove healing of surgical wounds, burn therapy to improve healingand/or reduce and minimize scarring, improving circulation within theskin, and the like.

The present disclosure can also be useful in improving wound healing,including but not limited to chronic skin ulcers, diabetic ulcers,thermal burn injuries, viral ulcers or disorders, periodontal diseaseand other dental disease. The present disclosure can be useful intreating the pancreas in diabetes. The present disclosure can be usefulfor in vitro fertilization enhancement, and the like. The presentdisclosure, in certain embodiments, is also useful in enhancing theeffects of devices that create an injury or wound in the process ofperforming cosmetic surgery including non-ablative thermal woundingtechniques for treating skin wrinkles, scars, stretch marks and otherskin disorders. Under such circumstances, it may be preferable to useconventional non-ablative thermal treatments in combination with themethods of the present disclosure. The instant application, in certainembodiments, are used in conjunction with micro- or surface abrasion,dermabrasion, or enzymatic or chemical peeling of the skin or topicalcosmeceutical applications, with or without nanoparticle application toenhance treatment, as the removal of the stratum corneum (and possiblyadditional epithelial layers) can prove beneficial for some treatmentregimen. The methods of the present disclosure are particularlyapplicable to, but are not limited to, acne treatment, hair removal,hair growth/hair follicle stimulation, reduction/prevention of malignantand non-malignant skin tumors, and skin rejuvenation, as describedherein.

The dermatologically therapeutic methods described herein may be formedusing nanoparticle irradiation alone, nanoparticle irradiation incombination with nano- or microparticles, or nanoparticle irradiationwith a composition comprising nano- or microparticles and one or moretherapeutic agents. Such nanoparticle irradiation may be produced by anyknown nanoparticle generator, and is preferably a focused nanoparticlegenerator capable of generating and irradiating focused nanoparticlewaves. Additionally, nanoparticle waves can be focused in tissues toprovide damage to local areas with a desirable size and shape.

Examples Example 1 Generation of Plasmonic Nanoparticles forThermomodulation

Plasmonic nanoparticles, including nanorods, hollow nanoshells, siliconnanoshells, nanoplates, nanorice, nanowires, nanopyramids, nanoprisms,nanoplates and other configurations described herein and known to thoseskilled in the art, are generated in size ranges from 1-1000 nm underconditions such that surface properties that facilitate deep follicularpenetration. Surface properties can be varied on one or multiple (2, 3,or 4) different dimensions to increase nanoparticle concentration in atarget tissue domain. Penetration into follicular openings of 10-200 umcan be maximized using the nanoparticles described herein. Here,nanoparticles sized in the range of about 10 to about 100 nm aregenerated, and are preferably assembled or formulated intomultiparticular structures having a size in the range of 100-300 nm.Alternatively, a coating (e.g., silica) is grown on uniparticularstructures to increase the particle size to the range of 100-300 nm ormore.

Surface-Modified Plasmonic Nanoparticles.

An exemplary preparation of surface-modified plasmonic nanoparticles isprovided as follows. Plasmonic nanoparticles are synthesized with stablecetryltrimethylamonium bromide (CTAB) coating and concentrated from anoptical density of 1 O.D. to 100, 200, 300, 400, or 500 O.D. through oneto three cycles of centrifugation at 16,000 rcf, with supernatantdecanting. Alternatively, CTAB-coated nanoparticles are concentrated andresuspended in 250 Amol/L 5-kDa methyl-polyethylene glycol (PEG)-thiolto make PEG-coated nanoparticles. Verification that PEG polymer stocksare fully reduced is performed using spectrophotometry to measure thethiol activity of polymer-thiols with 5,5-dithiobis(2-nitrobenzoic acid)against a DTT gradient. The solution of methy-PEG-thiol and CTAB-coatednanoparticles is mixed at room temperature for 1 h then dialyzed against5 kDa MWCO in 4 L distilled water for 24 h. Dialyzed samples areprocessed through 100-kDa filters to remove excess polymer.Quantification of the number of PEG polymers per particle is performedby surface-modifying nanoparticles with amino-PEG-thiol polymer andquantifying the number of amines with an SPDP assay. For testformulations, 100 O.D. solutions of CTAB-coated plasmonic nanoparticlesare made in distilled water, and 100 O.D. PEG-coated plasmonicnanoparticles are made in distilled water, ethanol, DMSO, or mineraloil. Plasmonic nanoparticles with silica shells are created by reactingnanoparticles with silicates such as tetra-ethyl-ortho-silicate (TEOS),sodium silicate, aminopropyletriethoxysilane (APTS), etc. to thicknessesof 5-50 nm or more. Control, vehicle-only formulations contain nonanoparticles.

Embedded Nanoparticles.

Nanoparticles are embedded (or encapsulated) in materials, which allowsfor the generation of a diverse range of sizes to tune their size.Particle sizes in the range of 100-2000 nm or 200-2000 nm have beenshown to enter the hair follicle without penetrating the dermis.Nanoparticles are encapsulated in silica, a syntheticpolylactide/co-glycolide co-polymer, porous lauryllactame/caprolactamnylon co-polymer, hydroxyethylcellulose, polyelectrolyte monolayers, oralternatively, in natural hydrogels such as hyaluronic acid, withoutsignificantly altering plasmon resonance properties. Nanoparticles areembedded within 100-2000 nm materials or 200-2000 nm materials withoutcovalent attachment or by cross-linking of amines, carboxyls or othermoieties on the nanoparticle surface to the polymer structure. Thesurface of the 100-2000 nm material or 200-2000 nm material may bemodified for an optimal zeta potential, hydrophilicity/hydrophobicity,and/or adsorption layer through techniques described herein.Furthermore, the shape of the aspect ratio of the polymer can bemodified from low to high to increase concentrations and depths ofpenetration of the embedded plasmonic nanoparticles. The nanoparticlesadvantageously have an aspect ratio greater than about 1.

Example 2 Formulation of Thermoablative Plasmonic Nanoparticles forTopical Delivery

Nanoparticles are generated as in Example 1 using an appropriate solvent(e.g., water, ethanol, dimethyl sulfoxide). The mixture comprising aplurality of nanoparticles in water is concentrated to about 100-500O.D. and exchanged for a new solvent by liquid chromatography, a solventexchange system, a centrifuge, precipitation, or dialysis. The solventmay include an alcohol (e.g., n-Butanol, isopropanol, n-Propanol,Ethanol, Methanol), a hydrocarbon (e.g., pentane, cyclopentane, hexane,cyclohexane, benzene, toluene, 1,4-Dioxane), chloroform, Diethyl-ether,water, an acid (e.g., acetic acid, formic acid), a base, acetone,dimethyl sulfoxide (DMSO), dimethylformamide (DMF), acetonitrile (MeCN),tetrahydrofuran (THF), dichloromethane (DCM) or ethylacetate. The newsolvent is combined with a cosmetically or pharmaceutically acceptablecarrier, thereby forming a nanoparticle composition. Generally, theparticles and carrier will form an emulsion.

Plasmonic nanoparticle formulations are provided that amplify orexpedite the penetration of nanoparticles into hair follicles. In someembodiments, nano- and micro-emulsions facilitate partitioning withinlipid-rich skin compartments such as the hair follicle. In someembodiments, nanoparticles are formulated in compositions containing0.5-2% v/v surfactants to enable disruption of the epidermal skinbarrier, emulsification of sebum, and improved mixing of hydrophilicnanoparticles in hydrophobic solutions or targeting to hydrophobic spacein the skin (e.g. between the hair shaft and surrounding follicle).Formulations of nanoparticles are also provided at variousconcentrations (1-20% w/v) in aqueous solutions, silicone/oil solvents,polypropylene gel, propylene glycol or creams (e.g. containing alcohols,oils, paraffins, colloidal silicas). In some embodiments,light-absorbing nanoparticles are utilized in solutions having tailoredpH, temperature, osmolyte concentration, viscosity, volatility, andother characteristics to improve light-absorbing nanoparticle entry intohair follicles.

Formulations are prepared to maximize nanoparticle stability (degree ofaggregation in solution), nanoparticle concentration, and nanoparticleabsorbance (degree of laser-induced heating at differentconcentrations).

When formulations of plasmonic nanoparticles are illuminated with aclinical laser with a wavelength coincident to the peak absorptionwavelength of the particle, the formulation heats to thermoablativetemperatures more rapidly and to a greater degree than conventionalclinical absorptive dyes. FIG. 2 compares the temperature profile ofplasmonic particles (1020 nm peak absorption wavelength) to conventionalclinical dyes carbon lotion, meladine spray and indocyanine green afterexposure to 1064 nm, 20 J/cm², 55 ms laser pulses. The temperatureincrease caused by pulsed 1064 nm laser light was more than 2.5 timesgreater for the plasmonic solution, compared to conventional clinicaldyes used at the same dilution (1:1000 dilution from clinicalconcentration, where clinical concentrations are as follows: carbon20-200 mg/ml, meladine 1 mg/ml, indocyanine green 5 mg/ml).

Example 3 Use of Plasmonic Nanoparticles for Thermomodulation of Hair

Individuals having blonde, red, gray, or lightly-colored hair are notadequately treated with existing light-based hair removal techniques.Provided herein are methods for using the compositions described hereinfor the selective removal or reduction of untreated blonde, red, gray,or lightly-colored hair. Plasmonic nanoparticles generated andformulated as described above are introduced into a target tissueregion, generally a skin region, and activated with laser-based hairremoval systems as known in the art in order to achieve effective hairremoval.

To achieve maximal penetration depth and concentration of plasmonicnanoparticles in the hair follicle and/or near components of thesebaceous gland including the sebaceous duct, the sebum, the epitheliallinking of the sebaceous gland, and/or near the bulge region includingthe stem cells, stem cell niche, epithelial lining of the bulge region,and/or near the follicular bulb, an optimal particle size of 30-800 nm(e.g., 100-800 nm) containing one or several plasmonic nanoparticles isconstructed. Nanoparticles encapsulating plasmonic nanoparticles can beformulated from any number of polymers or matrices. In some embodiments,the formulation contains a degradable or non-degradable polymer, e.g.,synthetic polylactide/co-glycolide co-polymer, porouslauryllactame/caprolactame nylon co-polymer, hydroxyethylcellulose,polyelectrolyte monolayers, or alternatively, in natural hydrogels suchas hyaluronic acid, gelatin and others. In further embodiments, ahydrogel PLGA, PEG-acrylate is included in the formulation.Preferentially, a matrix component such as silica, polystyrene orpolyethylene glycol is provided in the formulation to improve particlestability and enable facile removal from the skin surface afterapplication and follicle targeting. Other formulations include componentof surfactants (e.g. sodium dodecyl sulfate, sodium laureth 2-sulfate,ammonium lauryl sulfate, sodium octech-1/deceth-1 sulfate), a lipidbilayer, a liposome, or a microsome. Plasmonic nanoparticles includingnanorods, nanoshells, nanospheres, nanoplates, or nanorice can beencapsulated within a the polymer or lipid-based nanoparticle or matrixor deposited on the particle surface. Alternatively, nanoparticles inthe size range of 100-250 nm, 250-500 nm, 800 nm-1500 nm, or greaterthan 1500 nm can be used.

Pre-treatment of skin with mechanical or chemical exfoliation is used insome embodiments to remove hair-plugs and “open” the follicle forparticle delivery. Additionally, hairs can be shaven or waxed to createa void in the hair follicle for particles to fill. The use of physicalor thermal force amplifies or expedites the penetration of lightabsorbing nanoparticles and conjugates thereof into hair follicles, inpart by causing dilation of the hair follicle prior to application ofthe nanoparticles. For example, ultrasound and other sonic forces,mechanical vibrations, hair shaft manipulation (including pulling),physical force, thermal manipulation, and other treatments are utilizedto improve entry of light-absorbing nanoparticles into hair follicles.Nanoparticle formulation treatments are performed alone, in combination,sequentially or repeated 1-24 times.

An applicator is used to uniformly apply the composition ofnanoparticles into follicles. The applicator can be a sponge, a cloth,direct contact from a finger, a tube, a syringe, a device that appliessuction, an aerosol, a spray, or other means known in the art. In oneexample, a formulation of 1 ml of plasmonic nanoparticles at aconcentration of 100 O O.D. with peak resonance of 810 nm is applied toapproximately 200 cm² area of the skin of an adult human subject with asyringe. A cloth is used to evenly distribute solution across the skinarea and into the hair follicles. Deep massage from a mechanicalvibrator for 2 minutes with or without 1 MHz ultrasound for 5 minutes,is applied to drive particles deep into the follicle. Particlespenetrate 50-75% down the full length of the hair shaft atconcentrations sufficient to heat skin in a 100 μm radius at incrementaltemperatures of 5-20-fold greater than is generated in similar volumesof adjacent skin when irradiated by a Diode (810 nm) laser. Acetone,ethanol, or a debriding agent can be used to remove all particles fromthe surface of the skin that have not deposited in the follicle, inorder to reduced or prevent non-follicular heating of the skin.

Nanoparticle formulations are tested in ex vivo animal samples, ex vivohuman skin samples, and in vivo human skin including the assessmentof: 1) depth of nanoparticle penetration into hair follicles; 2)particle concentration achieved; 3) degree of heating achieved atdelivered nanoparticle concentrations; and 4) efficacy of photothermaldestruction including temporary and permanent hair removal, 5) clearanceof nanoparticles after treatment. To assess nanoparticle penetrationdepths, plasmonic nanoparticles surface-functionalized with fluorescentmolecules are visualized by fluorescence microscopy after histologicalsectioning or follicular biopsy (removal of hair shaft). Alternatively,plasmonic nanoparticles are directly visualized by dark field microscopyafter histological sectioning or follicular biopsy. To assessnanoparticle concentrations at various depths along the follicle,excised skin samples are separated by tape stripping or heat-basedtechniques, samples are dissolved for bulk analysis of metalconcentration by ICP-MS (inductively coupled plasma-mass spectrometry).The macroscopic degree of heating is validated by infrared thermographyof skin samples, and by assessment of skin sections subject to laserexposure for thermal damage markers. Finally, one can measure efficacyof photothermal destruction at the nanoparticle accumulation site byanalyzing histological cellular lesions at the target site, includingthe follicular hair shaft, inner root sheath, outer room sheath, andbulge region containing the stem cell niche, which contains the stemcells that contribute to new hair growth. As the bulge region isgenerally localized about midway (˜50% down the length of) the hairshaft, permanent hair removal is sufficiently achieved by accumulationof plasmonic nanoparticles to this depth. In some situations,nanoparticle delivery may also generate a heat gradient emitting furtherdown the hair shaft. Animal studies are useful to demonstrate theefficacy of unpigmented hair removal by comparing heat profiles, thermalablation of hair shaft, and thermal damage of bulge stem cells intreated hairless rodents, albino rodents and dark-haired rodents.Efficacy on live human skin is measured by measuring hair counts at 3and 12 month follow ups. Biopsies are taken from select patients at 2,4, and 6 week follow ups to verify that nanoparticles are cleared fromthe skin without embedding in the dermis.

Hair Follicle Penetration of Fluorescently-Labeled NanoparticlesDetermined Using Porcine Skin Explants and Confocal Imaging.

A 25 mg/ml aqueous solution silicon dioxide-coated nanoparticles (200 nmdiameter) was contacted with freshly thawed porcine skin, after whichexcess nanoparticle suspension was removed and manual massage wasperformed for three minutes. The explant was sectioned and subjected toconfocal imaging. As shown in FIG. 3A, explant sections were imaged atangles to the hair follicles in 60 μm planes; Plane 1 shows the follicleinfundibulum, while Plane 2 shows the distal regions of the follicle.FIG. 3B demonstrates representative confocal images showing that rednanoparticles (548 nm absorbance) are visible within both thesuperficial and deep follicles, but are not detectable in dermal layersbeneath the follicles. FIG. 3C shows high-magnification imaging of rednanoparticles localized to and retained within a deep follicle (˜400μm). Green color indicates tissue autofluorescence (488 nm).

Hair Follicle Penetration of Plasmonic Nanoparticles Determined UsingPorcine Skin and Dark Field Imaging.

A 100 O.D. suspension of plasmonic nanoparticles (200 nm diameter) wascontacted with freshly thawed porcine skin, after which excessnanoparticle suspension was removed and manual massage performed forthree minutes. The procedure was repeated for a total of 3 applications,and surface residue removed with several 3-5 applications of alternatingwater and ethanol. The skin sample was excised, fixed, sectioned alonghorizontal plane and subjected to dark field imaging. As shown in FIG.4A, skin samples were sectioned and imaged horizontal to the hairfollicle at various depths. In skin section images, plasmonicnanoparticles were observed as bright blue color point sources at depthsup to 1.2 mm deep in porcine follicle spaces (FIG. 4B). Control sampleswith no plasmonic nanoparticles were clearly differentiated (FIG. 4C).ICP-MS is also performed on skin sections to assess nanoparticleconcentrations at various depths along the follicle.

Hair Follicle Penetration of Nanoparticles in Hairless Rodents, AlbinoRodents and Dark-Haired Rodents.

White-haired Swiss Webster mice (n=3) at 8 weeks old are anesthetizedwith injectable ketamine/xylazine anesthetic solution and dorsal backskin and hair washed and dried. Prior to formulation administration,three 10 cm×10 cm areas are demarcated by permanent marker on each mouseand subjected to hair removal by 1) electric razor, 2) Nair depilationreagent, or 3) warm wax/rosin mixture application and stripping. Eachmouse is treated by pipette with up to 3 nanoparticle formulations, inquadruplicate 5-μl spot sizes per demarcated skin area (up to 12 spotsper area or 36 spots per mouse). Precise spot locations are demarcatedwith pen prior to pipetting. Duplicate treatment spots on the dorsalleft side are massaged into skin for 5 minutes, while duplicatetreatment spots on the dorsal right side are applied without massage.Thirty minutes after application, mice are sacrificed by carbon dioxideasphyxiation and cervical dislocation, and skin is carefully excised andpunched into sections along spot size demarcations. Skin biopsies arefixed in 10% paraformaldehyde, paraffin-embedded, and cut into 5-umsections on a microtome in transverse directions. Slides with mountedparaffin sections are deparaffinized and stained with hematoxylin andeosin (H&E) or kept unstained for dark field microscopy. Using H&Estaining, light microscopy and/or dark field microscopy, greater than 50follicles per formulation are imaged, and scoring is performed for skinsections for visible macroscopic nanoparticle accumulation in thefollicle, along the hair shaft, at the site of the putative bulge stemcell niche, and at the depth of the follicle bulb. On serialhistological sections, a silver enhancement staining kit based on sodiumthiosulfate may be used to enlarge the plasmonic nanoparticle signal viathe precipitation of metallic silver. Phase and dark field micrographsare captured and used to record the depths of follicular penetration foreach nanoparticle formulation and method of application. ICP-MS is alsoperformed on skin sections to assess nanoparticle concentrations atvarious depths along the follicle.

Assessment of Photothermal Destruction at the Nanoparticle AccumulationSite.

Treated areas of pig, human or mouse skin are irradiated with a lasercoincident with the peak absorption wavelength of nanoparticles (e.g.1064 nm YAG laser for 1020 nm plasmonic particles) using clinicalparameters (1 s exposure of 30-50 J/cm² and a pulse width of 10-50 ms).To determine microscopic photothermal damage of target skin structuressuch as the hair follicle and hair follicle bulge stem cells, at tendays after application and irradiation, human subjects receive lidocaineinjections to numb treatment areas and skin is carefully excised andpunched into sections along spot size demarcations. Fresh human skinbiopsies or explanted human and animal skin samples are fixed in 10%paraformaldehyde, paraffin-embedded, and cut into 5-um sections on amicrotome in transverse directions, or they are fixed in Zamboni'ssolution with 2% picric acid and cryosectioned by freezing slidingmicrotome. Slides with mounted paraffin sections are deparaffinized andstained with hematoxylin and eosin (H&E). Histological sections areexamined at various depths for markers of thermal damage andinflammation. Hematoxylin and eosin (H&E) is used to image skin andfollicle microanatomy and indicate degeneration of hair shafts, atrophyof sebaceous glands, and cell vacuolization (indicating cellulardamage). Nitro blue tetrazolium chloride (NBTC), a lactate dehydrogenasestain that is lost upon thermal injury to cells, is used to assessdamage to keratinocytes. Cellular damage in follicles of skin samplesreceiving plasmonic nanoparticle plus laser treatment is scored andcompared to those receiving laser treatment alone. Live treated humanskin areas are also followed clinically for 2 weeks to 3 monthsfollowing plasmonic nanoparticle+laser treatment, or during repeatedplasmonic nanoparticle+laser treatments, and compared to baselinedigital photograph taken prior to first treatment, and to negativecontrol laser only treatments. Clinical observations of hair removal, aswell as erythema, edema, discomfort, irritation or scarring, are notedto determine degree of non-specific thermal damage.

Effect of Plasmonic Particle Coating on Specificity of Delivery andPhotothermal Heating.

Preferentially, a matrix component such as silica, polystyrene orpolyethylene glycol is provided in the formulation to improve particlestability and enable facile removal from the skin surface afterapplication and follicle targeting. Acetone, ethanol, or a debridingagent can be used to remove all particles from the surface of the skinthat have not deposited in the follicle, in order to reduced or preventnon-follicular heating of the skin. In FIG. 5, live human skin wastreated with Uncoated plasmonic particles compared to Silica-coatedplasmonic particles, prior to laser-irradiation and comparison to noparticle treatment (laser only) controls. Pre-treatment of skin,including shaving with razor and microdermabrasion (15 sec, mediumsetting) to remove hair-plugs and “open” the follicle for particledelivery, was performed on both forearms. Human forearm skin wasirradiated with 810 nm laser pulses (30 J/cm², 30 ms, 2 passes) alone(FIG. 5A), or after treatment with a formulation of 830 nm resonant,Uncoated plasmonic nanoparticles in 20% propylene glycol (FIG. 5B). Theplasmonic nanoparticle formulation was applied with 3 minute massage andrepeated 3 times, and the skin surface wiped with 3 applications ofalternative water and ethanol before laser irradiation. At 30 minutesfollowing laser irradiation, non-specific clinical burns were observeddue to significant photothermal heating of residual, Uncoated particleson the skin surface (FIG. 5B). Live human skin was also irradiated with1064 nm laser pulses (40 J/cm², 55 ms, 3 passes) alone (FIG. 5C), orafter treatment with a formulation of 1020 nm resonant, Silica-coatedplasmonic nanoparticles in 20% propylene glycol (FIG. 5D). The plasmonicnanoparticle formulation was applied with 3 minute massage and repeated3 times, and the skin surface wiped with 3 applications of alternativewater and ethanol before laser irradiation. At 30 minutes followinglaser irradiation, no evidence of burning of the skin or erythema wasobserved, as Silica-coated particles could be sufficiently wiped fromthe skin surface (FIG. 5D). Magnified photography of the skin areatreated with Silica-coated particles+Laser shows specific photothermaldamage (perifollicular erythema and edema) in the nanoparticle-targetedsite, without damage to surrounding or non-particle-treated tissues(FIG. 6).

Example 4 Use of Plasmonic Nanoparticles for Acne Treatment

Provided herein are methods for using the compositions described hereinfor the treatment of acne vulgaris and other acnes and acne-like skinconditions, but the selective targeting of sebaceous follicles,particularly the sebaceous glands and/or hair follicles. Plasmonicnanoparticles generated and formulated as described above are introducedinto a target tissue region, generally a skin region, and activated withlaser-based systems as known in the art in order to achieve effectivehair removal.

To achieve maximal penetration depth and concentration of plasmonicnanoparticles in the hair follicle and/or near components of thesebaceous gland including the sebaceous duct, the sebum, the epitheliallinking of the sebaceous gland, and/or near the bulge region includingthe stem cells, stem cell niche, epithelial lining of the bulge region,and/or near the follicular bulb, an optimal particle size of 100-800 nmcontaining one or several plasmonic nanoparticles is constructed.Nanoparticles encapsulating plasmonic nanoparticles can be formulatedfrom any number of polymers or matrices. In some embodiments, theformulation contains a degradable or non-degradable polymer, e.g.,synthetic polylactide/co-glycolide co-polymer, porouslauryllactame/caprolactame nylon co-polymer, hydroxyethylcellulose,polyelectrolyte monolayers, or alternatively, in natural hydrogels suchas hyaluronic acid, gelatin and others. In further embodiments, ahydrogel PLGA, PEG-acrylate is included in the formulation.Preferentially, a matrix component such as silica, polystyrene orpolyethylene glycol is provided in the formulation to improve particlestability and enable facile removal from the skin surface afterapplication and follicle targeting. Preferentially, formulations includesurfactants (e.g. sodium dodecyl sulfate, sodium laureth 2-sulfate,ammonium lauryl sulfate, sodium octech-1/deceth-1 sulfate), componentsof a lipid bilayer, a liposome, or a microsome. Surfactants disrupt theepidermal skin barrier, emulsify sebum, improve mixing of hydrophilicnanoparticles with hydrophobic solutions, and reduce entropic barriersto delivering hydrophilic particles to hydrophobic regions of the skin(e.g. between the hair shaft and surrounding sheath or follicle).Plasmonic nanoparticles including nanorods, nanoshells, nanospheres, ornanorice can be encapsulated within the polymer nanoparticle or matrixor deposited on the particle surface. Alternatively, nanoparticles inthe size range of 100-250 nm, 250-500 nm, 800 nm-1500 nm, or greaterthan 1500 nm can be used.

The use of physical or thermal force amplifies or expedites thepenetration of light absorbing nanoparticles and conjugates thereof intohair follicles and/or sebaceous glands, in part by causing dilation ofthe hair follicle prior to application of the nanoparticles. Forexample, ultrasound and other sonic forces, mechanical vibrations, hairshaft manipulation (including pulling), physical force, thermalmanipulation, and other treatments are utilized to improve entry oflight-absorbing nanoparticles into hair follicles and/or sebaceousglands. Nanoparticle formulation treatments are performed alone, incombination, sequentially or repeated 1-24 times.

Prior to application of the plasmonic nanoparticles, a pre-treatmentstep of removing excess sebum from the surface of the skin may beperformed using chemical and/or mechanical means. Pre-treatment of skinwith mechanical or chemical exfoliation is used in some embodiments toremove hair-plugs and “open” the follicle for particle delivery.Additionally, hairs can be shaven or waxed to create a void in the hairfollicle for particles to fill.

An applicator is used to uniformly apply the composition ofnanoparticles into follicles. The applicator can be a sponge, a cloth,direct contact from a finger, a tube, a syringe, a device that appliessuction, an aerosol, a spray, or other means known in the art. In oneexample, a formulation of 1 ml of plasmonic nanoparticles at aconcentration of 100 O.D. with peak resonance of 810 nm is applied toapproximately 200 cm² area of the skin of an adult human subject with asyringe. A cloth is used to evenly distribute solution across the skinarea and into the hair follicles. Massage from a mechanical vibrator for2 minutes with or without ultrasound at 1 MHz for 5 minutes is appliedto drive particles deep into the follicle. Particles penetrate ˜50% downthe full length of the hair shaft at concentrations sufficient to heatskin in a 100 um radius at incremental temperatures of 5-20-fold greaterthan is generated in similar volumes of adjacent skin when irradiated bya Diode (810 nm) laser. Acetone, ethanol, or a debriding agent can beused to remove all particles from the surface of the skin that have notdeposited in the follicle, in order to reduced or prevent non-follicularheating of the skin.

Delivery of Plasmonic Nanoparticles to the Sebaceous Gland DeterminedUsing Human Abdominoplasty Skin and Dark Field Imaging.

The human sebaceous gland exists within the pilosebaceous unitconsisting of the hair, hair follicle, arrector pili muscle andsebaceous gland. In FIG. 7A, a human skin biopsy is immunostained withantibodies against Collagen IV (basement membrane marker, blue) and PGP9.5 (nerve marker, green) to visualize representative pilosebaceous unitmicroanatomy, including the hair follicle (HF), sebaceous gland (SG) andarrector pili muscle. To deliver nanoparticles to the hair follicle andsebaceous gland, skin was first pre-treated with shaving to removeextruding hair, microdermabrasion (15 sec, medium setting) to removehair-plugs and corneocytes, and chemical depilation to “open” folliclemicrowells for particle delivery. A 100 O.D. suspension of plasmonicnanoparticles (200 nm diameter), formulated in 1% sodium dodecyl sulfate(SDS) and 20% propylene glycol (PG) was contacted with excised humanabdominoplasty skin, after which excess nanoparticle suspension wasremoved and manual massage performed for three minutes, followed byultrasound (1 MHz) for 5 minutes. The procedure was repeated for a totalof 3 applications, and surface residue removed with 3-5 applications ofalternating water and ethanol. The skin sample was excised, fixed,sectioned along horizontal planes and subjected to dark field imaging.As assessed by dark field imaging of horizontal skin sections,compositions of plasmonic nanoparticles with a cosmetically acceptablecarrier of 1% SDS/20% PG administered with massage and ultrasound can bedelivered 400-600 μm deep into the human follicle and specifically intothe sebaceous gland (FIG. 7B).

Cosmetic Formulations for Follicle and Sebaceous Gland Delivery in HumanSkin.

Preferentially, formulations include surfactants (e.g. sodium dodecylsulfate, sodium laureth 2-sulfate, ammonium lauryl sulfate, sodiumoctech-1/deceth-1 sulfate), components of a lipid bilayer, a liposome,or a microsome. Surfactants disrupt the epidermal skin barrier andemulsify the sebum to enable improved mixing of hydrophilicnanoparticles in hydrophobic solutions. Humectants such as propyleneglycol are used to help improve topical viscosity and maintainphysiological pH. To demonstrate the efficacy and mechanism of exemplarycosmetic formulations for human sebaceous gland delivery, skin was firstpre-treated with shaving to remove extruding hair, micro dermabrasion(15 sec, medium setting) to remove hair-plugs and corneocytes, andchemical depilation to “open” follicle microwells for particle delivery.Two separate 100 O.D. suspensions of plasmonic nanoparticles (200 nmdiameter) were formulated in 1% sodium dodecyl sulfate and 20% propyleneglycol (SDS/PG) or in 1% sodium laureth-2-sulfate and 20% propyleneglycol (SLES/PG). Formulations were contacted with two separate excisedhuman abdominoplasty skin samples, and massage for 3 minutes followed byultrasound (1 MHz) for 5 min was performed to drive particles deep intothe follicles. The procedure was repeated for a total of 3 applications,and surface residue removed with 3-5 applications of alternating waterand ethanol. The skin sample was excised, fixed, sectioned alonghorizontal planes and subjected to dark field imaging to assess particledelivery. As assessed by dark field imaging of horizontal skin sections,compositions of plasmonic nanoparticles with a cosmetically acceptablecarrier of 1% SLES/20% administered with massage and ultrasound can bedelivered 400-600 μm deep into the human follicle and specifically intothe sebaceous gland (FIG. 8B).

Impact of Massage Vs. Ultrasound on Nanoparticle Delivery to HumanFollicles and Sebaceous Gland.

Ultrasound and other sonic forces, mechanical vibrations, hair shaftmanipulation (including pulling), physical force, thermal manipulation,and other treatments are utilized to improve entry of light-absorbingnanoparticles into hair follicles and/or sebaceous glands. Mechanicalmassage improves follicular penetration through hair shaft ‘pumping’mechanisms, while ultrasound enhances transdermal drug delivery throughtemporary disruption of the skin's lipid bilayer, bubble formation, andliquid microstreaming. To characterize the effects of massage decoupledfrom ultrasound, skin was first pre-treated with shaving to removeextruding hair, micro dermabrasion (15 sec, medium setting) to removehair-plugs and corneocytes, and chemical depilation to “open” folliclemicrowells for particle delivery. A 100 O.D. suspension of plasmonicnanoparticles (200 nm diameter), formulated in 1% sodium dodecyl sulfate(SDS) and 20% propylene glycol (PG), was contacted with three separateexcised human abdominoplasty skin samples. In the three treated humanskin samples, massage only was performed for 3 minutes, ultrasound only(1 MHz) was performed for 5 minutes, or massage followed by ultrasoundwas performed to drive particles deep into the follicles. In a fourthsample, no particles were applied to skin. The procedure was repeatedfor a total of 3 applications, and surface residue removed with 3-5applications of alternating water and ethanol. The skin sample wasexcised, fixed, sectioned along horizontal planes and subjected to darkfield imaging to assess particle delivery. As assessed by dark fieldimaging of horizontal skin sections, compositions of plasmonicnanoparticles with a cosmetically acceptable carrier of 1% SLES/20%administered via ultrasound deliver more plasmonic nanoparticles to theinfundibulum versus massage, albeit both mechanisms facilitate delivery(FIG. 9).

Additional Plasmonic Nanoparticle Formulations for Follicle andSebaceous Land Delivery in Human Skin.

In some embodiments, plasmonic nanoparticles include nanorods,nanoshells, nanospheres, or nanorice, or plasmonic nanoparticlesencapsulated within the polymer nanoparticle or matrix or deposited onthe particle surface. Preferentially, a matrix component such as silica,polystyrene or polyethylene glycol is provided in the formulation toimprove particle stability and enable facile removal from the skinsurface after application and follicle targeting. To demonstrate theformulation of additional plasmonic nanoparticle shapes andconcentrations for follicle, infundibulum, and sebaceous gland delivery,skin was first pre-treated with shaving to remove extruding hair,microdermabrasion (15 sec, medium setting) to remove hair-plugs andcorneocytes, and chemical depilation to “open” follicle microwells forparticle delivery. Separately, 10 O.D. suspensions of Silica-coatednanoplates, 30 O.D. suspensions of polyethylene-glycol coated plasmonicnanorods, and fluorescent silica particles were formulated in 1% sodiumdodecyl sulfate and 20% propylene glycol. Formulations were contactedwith three separate excised human abdominoplasty skin samples, andmassage for 3 minutes followed by ultrasound (1 MHz) for 5 min wasperformed to drive particles deep into the follicles. The procedure wasrepeated for a total of 3 applications, and surface residue removed with3-5 applications of alternating water and ethanol. The skin sample wasexcised, fixed, sectioned along horizontal planes and subjected to darkfield imaging to assess particle delivery. As assessed by dark fieldimaging of horizontal skin sections, compositions of Polyethylene glycol(PEG)-coated nanorods (gold, 15×30 nm dimension) in cosmeticallyacceptable carrier, administered via ultrasound and massage, wereobserved within the follicle infundibulum at 200 um deep (FIG. 10A).Compositions of plasmonic nanoparticles (Silica-coated nanoplates) atlower concentration (10 O.D.), were apparent at 400-600 um deep in thefollicle and in the sebaceous gland (open arrow), albeit at lowerconcentration than comparable particles in a similar cosmetic carrier at100 O.D (FIG. 10B).

Assessment of Photothermal Destruction of Sebaceous Gland and TargetedSkin Structures.

Nanoparticle formulations are tested in ex vivo animal skin samples, exvivo human skin samples, and in vivo human skin as described in Example3. One can measure efficacy of photothermal destruction at thenanoparticle accumulation site by measuring thermal damage to sebocytesand reduction in sebum production in the treated sebaceous follicles. Toassess photothermal destruction, human skin is first pre-treated withshaving to remove extruding hair, micro dermabrasion (15 sec, mediumsetting) to remove hair-plugs and corneocytes, and chemical depilationto “open” follicle microwells for particle delivery. Skin is contactedwith a 100 O.D. suspension of 810 nm resonant plasmonic nanoparticles(200 nm diameter), and is massaged for 3 minutes followed by ultrasound(1 MHz) for 5 min to drive particles deep into the follicles. Theprocedure is repeated for a total of 3 applications, and surface residueremoved with 3-5 applications of alternating water and ethanol. Treatedhuman skin samples are laser irradiated with 810 nm laser (40 J/cm², 30ms, 5 pulses), and compared to laser only treated human skin. Human skinis biopsied, fixed in Zamboni's solution with 2% picric acid, andcryosectioned by freezing sliding microtome. Slides with mountedparaffin sections are deparaffinized and stained with hematoxylin andeosin (H&E). Histological sections are examined at various depths formarkers of thermal damage and inflammation. Hematoxylin and eosin (H&E)is used to image skin and follicle microanatomy and indicatedegeneration of hair shafts, atrophy of sebaceous glands, and cellvacuolization (indicating cellular damage). Nitro blue tetrazoliumchloride (NBTC), a lactate dehydrogenase stain that is lost upon thermalinjury to cells, may also be used to assess damage to keratinocytes vs.sebocytes. An intracellular stain, Oil-Red-O, may be used to determinelipid and sebum oil content in treated samples. Sebum excretion ratesare measured on in vivo skin at 1-3 months follow up usingsebum-absorbant tapes to demonstrate functional change in sebum flow.Clearance and prevention of acne lesions is measured by patient reportedoutcomes and counting acne lesions at 1-3 months follow up.

Example 5 Formulation of Thermoablative Plasmonic Nanoparticles forVascular Ablation

Formulations are prepared to maximize nanoparticle stability (degree ofaggregation in solution), nanoparticle concentration, and nanoparticleabsorbance (degree of laser-induced heating at different concentrations)once injected into the blood stream. Nanoparticles are generated as inExample 1 using an appropriate solvent. The mixture comprising aplurality of nanoparticles in water is concentrated to about 100-500 ODat peak absorbance and exchanged for a new solvent by liquidchromatography, a solvent exchange system, a centrifuge, precipitation,or dialysis. Typical exchange solvent is 0.15 mol/L NaCl, 0.1 mol/L Naphosphate buffer (pH 7.2).

Example 6 Use of Plasmonic Nanoparticles for Thermoablation ofComponent(s) of Vessels and Microvessels

Nanoparticle-containing compositions are administered, typicallyintravascularly. Subsequent to such administration of plasmonicnanoparticles, a laser matched to the peak plasmonic resonance of theparticles (e.g., 755 nm, 810 nm, or 1064 nm) is applied to heatnanoparticles and surrounding tissue. Pulse widths of 10-100 ns, 100ns-1 ms, 1-10 ms, 10-100 ms, 100-1000 ms or continuous wave irradiationis used to achieve thermal heat gradients and localized heating in thevicinity of particle or particles of 20-200 nm. 200 nm-2 μm, 2-20 μm,20-200 μm, 200 μm-2 mm. Thermal gradients of 20-200 nm are achieved fromindividual particles. Supra millimeter thermal gradients are achieved bythe collective heat deposition of many particles in veins with diametersof several hundred microns or more. Irradiation is applied from 1 pulseto many pulses over seconds to minutes. A cooling device for epidermallayers is used concomitant to irradiation to reduce pain and preventthermal damage elsewhere. Laser position, fluence, wavelength, angle ofincidence, pattern of irradiation is modified to achieve irradiation ofvessels at specific depths between 0-10 mm, while avoiding heating ofnon-target vasculature. Alternatively, laser or light is administeredthrough fiber optic waveguide administered via a catheter to heat theparticles in larger veins.

In one embodiment a flank of the tissue is irradiated with 2 W/cm², 810nm, 1 cm beam diameter after injection of PEG-nanorods with peak plasmonresonance at 810 nm. Thermographic imaging is used to assess surfacetemperature of tissue immediately after irradiation.

Assessment of thermal damage to component(s) of vessels, microvessels,or capillaries. Thirty minutes after application, target vessels and thesurrounding supporting tissue (e.g. skin) are removed. Biopsies arefixed in 10% paraformaldehyde, paraffin-embedded, and cut into 5-umsections on a microtome in transverse directions. Slides with mountedparaffin sections are deparaffinized and stained with hematoxylin andeosin (H&E) or silver enhancement staining. Using H&E staining and lightmicroscopy, one or several vessels, microvessels, and capillaries can beimaged. Scoring is performed for visible thermal damage of the vesselstructures. Additionally, vessel staining (e.g. CD31 stain) is performedto clearly identify vascular structures within tissue samples.

Example 7 Determination of Efficiency of Conversion of Light to ThermalEnergy

A suspension of plasmonic nanoparticles (silica-coated nanoplates havinga diameter of about 100-200 nm, as described here) was prepared byformulating the plasmonic nanoparticles in 20% propylene glycol in waterto a concentration of about 1000 O.D., and the ability of thissuspension to convert laser light to thermal energy was determined.Available commercial and research products, e.g., stock solutions ofcarbon lotion (20-200 mg/ml carbon, TelsarSoftLight), Meladine spray (1mg/ml melanin, Creative Technologies), Indocyanine green (5 mg/ml inwater, Sigma Aldrich), and vehicle control (20% propylene glycol inwater) were also tested. All solutions were diluted 1:1 000 from theirindicated stock solution concentration, loaded at 90 μl per well into a96-well plate, and baseline temperatures were measured by K thermocouplewith micrometer (ExTech Instruments, Waltham Mass.) and recorded.Solutions were then irradiated with repeated laser pulses at variouswavelengths (e.g., 1064 nm, 810 nm, and 755 nm), fluence (e.g., 10, 20,and 30 J/cm2) and pulse sequence parameters (e.g., 30 ms and 55 ms).Following each sequential laser pulse, up to a total of 8 pulses,solution temperatures were measured and recorded. As shown in FIGS.11A-11B, a series of plasmonic nanoparticle (PNP) formulations (labeledSL-001 and SL-002) exhibited ultra-high absorption compared to existingcommercial and research chromophores. (FIGS. 11A, B) Rate of temperatureincrease over sequential laser pulses for PNP formulation SL-001 (FIG.11A, closed circle), resonant at 1064 nm laser wavelength, uponirradiation with 1064 nm laser (A), and SL-002 (FIG. 11B closed circle),resonant at 810 nm laser wavelength, upon irradiation with 810 nm laser(B). Control solutions are as follows: Carbon lotion (open triangle),Meladine spray (closed square), Indocyanine green (open diamond), and20% propylene glycol (closed triangle). All solutions were diluted1:1000 from stock clinical concentration for laser irradiation andtemperature measurements. For A, n=2 and error bars are s.d. of themean.

Example 8

Quantitation of nanoparticle delivery into target tissues. Redfluorescent nanoparticles (Corpuscular Inc., Cold Spring, N.Y.) werecontacted with isolated porcine skin explants as follows. A 2.5 mg/mlsolution of SiO₂, 200 nm diameter, 548 nm emission particles in 20%propylene glycol was pipetted onto the skin surface and mechanicallymassaged into the tissue explant. An ethanol wipe was used to removenon-penetrating particles. As shown in FIGS. 12A-12B, the providedformulations of nanoparticles (NPs) deeply and specifically penetrate exvivo porcine skin. FIG. 12A demonstrates representative surveyfluorescence image of porcine skin, treated with red fluorescent NPs andhistologically sectioned. Red (light contrast) NPs are imaged afterpenetrating the hair follicle infundibulum (arrows) and deep follicle,but not in the underlying dermis. FIG. 12B shows representative confocalimages show red NPs within superficial and deep follicle (−870/−tm) athigh and low magnification. Green (dark contrast) is tissueautofluorescence (488 nm emission). Scale bars as labeled 1 mm (A), 10μm (B, left), 50 μm (B, right).

Further, formulations of nanoparticles (NPs) with silica coating deeplyand specifically penetrate in vivo human skin. A region of an upper armof a male human subject having skin Type 3 was treated with the rednanoparticles essentially as described above. Shown in FIGS. 13A and 13Bare representative confocal images of biopsies taken from the invivo-treated human skin, which were sectioned and immunostained for skinmarkers. Left-‘TH 2A R med’ sample shows red hair follicle fluorescenceafter red NP application with massage, ultrasound, and no pre-depilationwith waxing; Middle ‘TH 2C L’ sample shows red hair folliclefluorescence after red NP application with massage, ultration, andpre-depilation with waxing; Right—‘TH 1A Control’ shows background redautofluorescence of hair follicle. FIG. 13A is 3 color image where redis NPs, blue is collagen IV (staining basement membrane) and green isPGP 9.5 (staining nerve fiber). FIG. 13B shows red channel only in blackand white. Scale bars as labeled 100 μm.

As will be understood by the skilled artisan, the subject matterdescribed herein may be embodied in other specific forms withoutdeparting from the spirit or essential characteristics thereof. Theforegoing embodiments are therefore to be considered in all respectsillustrative rather than limiting on the invention described herein.

What is claimed is:
 1. A method for performing thermoablation,comprising the steps of contacting a target tissue region of a mammaliansubject with a composition comprising a plurality of plasmonicnanoparticles under conditions such that an effective amount of theplasmonic nanoparticles localize to a domain of the target tissueregion; and exposing the target tissue region to energy delivered from anonlinear excitation surface plasmon resonance source in an amounteffective to induce thermoablation of the domain of the target tissueregion, wherein the nanoparticle comprises an optical absorption ofabout 200 nm to about 10,000 nm.